Crystalline 4-(3-chloro-2-fluoroanilino)-7 methoxy-6-...

ABSTRACT

4-(3-chloro-2-fluoroanilino)-7-methoxy-6-{[1-(N-methylcarbamoylmethyl)piperidin-4-yl]oxy}quinazoline difumarate, pharmaceutical compositions containing the difumarate, the use of the difumarate in the treatment of hyperproliferative disorders such as cancer and processes for the manufacture of the difumarate are described.

CROSS REFERENCE TO RELATED APPLICATIONS

The present application claims benefit under 35 U.S.C. §119(e) of U.S.Provisional Application Ser. No. 61/052,706 filed on May 13, 2008 andU.S. Provisional Application Ser. No. 61/110,637 filed on Nov. 3, 2008,the contents of which are herein incorporated by reference in itsentirety.

BACKGROUND

The erbB family of receptor tyrosine kinases, which include EGFR, erbB2,erbB3 and erbB4, are frequently involved in driving the proliferationand survival of tumour cells and as such the erbB family of receptors isimplicated in a number of epithelial cancers (reviewed in Olayioye etal., EMBO J., 2000, 19, 3159), including for example breast cancer(Sainsbury et al., Brit. J. Cancer, 1988, 58, 458; Guerin et al.,Oncogene Res., 1988, 3, 21; Slamon et al., Science, 1989, 244, 707;Klijn et al., Breast Cancer Res. Treat., 1994, 29, 73 and reviewed inSalomon et al., Crit. Rev. Oncol. Hematol., 1995, 19, 183), non-smallcell lung cancers (NSCLCs) including adenocarcinomas (Cerny et al.,Brit. J. Cancer, 1986, 54, 265; Reubi et al., Int. J. Cancer, 1990, 45,269; Rusch et al., Cancer Research, 1993, 53, 2379; Brabender et al,Clin. Cancer Res., 2001, 7, 1850) as well as other cancers of the lung(Hendler et al., Cancer Cells, 1989, 7, 347; Ohsaki et al., Oncol. Rep.,2000, 7, 603), bladder cancer (Neal et al., Lancet, 1985, 366; Chow etal., Clin. Cancer Res., 2001, 7, 1957, Zhau et al., Mol Carcinog., 3,254), oesophageal cancer (Mukaida et al., Cancer, 1991, 68, 142),gastrointestinal cancer such as colon, rectal or stomach cancer (Bolenet al., Oncogene Res., 1987, 1, 149; Kapitanovic et al.,Gastroenterology, 2000, 112, 1103; Ross et al., Cancer Invest., 2001,19, 554), cancer of the prostate (Visakorpi et al., Histochem. J., 1992,24, 481; Kumar et al., 2000, 32, 73; Scher et al., J. Natl. CancerInst., 2000, 92, 1866), leukaemia (Konaka et al, Cell, 1984, 37, 1035,Martin-Subero et al., Cancer Genet Cytogenet., 2001, 127, 174), ovarian(Hellstrom et al., Cancer Res., 2001, 61, 2420), head and neck (Shiga etal., Head Neck, 2000, 22, 599) or pancreatic cancer (Ovotny et al.,Neoplasma, 2001, 48, 188).

Accordingly it has been recognised that an inhibitor of erbB receptortyrosine kinases should be of value as a selective inhibitor of thegrowth of certain carcinomas. A number of erbB tyrosine kinaseinhibitors have demonstrated clinical benefit and a number of erbBtyrosine kinase inhibitors have been approved for use in the treatmentof cancer. For example, the EGFR tyrosine kinase inhibitors gefitiniband erlotinib for the treatment of advanced non-small cell lung cancerand lapatinib, which has erbB2 tyrosine kinase inhibitory activity, foruse in metastatic breast cancer. Several other EGFR and erbB2 tyrosinekinase inhibitors are currently in development.

Compound (I) is disclosed in International Patent ApplicationPublication number WO2005/028469 as Example 1 therein and is of thestructure:

Compound (I) is an erbB receptor tyrosine kinase inhibitor, inparticular Compound (I) is a potent inhibitor of EGFR and erbB2 receptortyrosine kinases.

There is a growing body of pre- and clinical evidence suggesting that,in addition to signalling via EGFR and erbB2 homodimers, cell signallingmediated by EGFR, erbB2 & erbB3 heterodimers may be an importantoncogenic signalling pathway (Sergina et al., Nature, 2007, 445, 437;Ritter et al., Clin Cancer Res. 2007, 13, 4909; Johnston et al., JCO,2008, 26, 1066). Since erbB3 does not have an intrinsic tyrosine kinaseactivity, activation of the erbB3 receptor is achieved only through theformation of heterodimeric receptor complexes with other kinase-activereceptors including particularly EGFR and erbB2. EGFR and erbB2heterodimers formed with erbB3 are thought to drive tumour growth intumours where these receptors are expressed.

We have found in pre-clinical experiments that Compound (I) alsoinhibits erbB3 mediated signalling through the inhibition ofphosphorylation of erbB3 following ligand stimulated EGFR/erbB3 and/orerbB2/erbB3 heterodimerisation. Accordingly, Compound (I) exhibits aunique erbB tyrosine kinase inhibitory effect compared to other erbBtyrosine kinase inhibitors such as gefitinib or erlotinib that actprimarily as EGFR tyrosine kinase inhibitors. We have carried outpre-clinical studies which suggest that Compound (I) exhibits improvedanti-tumour effects compared to EGFR tyrosine kinase inhibitors such asgefitinib and erlotinib. Without wishing to be bound by theory, it isthought that the improved properties may result from the inhibition ofthe erbB3 mediated signalling by Compound (I).

WO2005/028469 indicates that the compounds disclosed therein may beprepared in the form of a pharmaceutically acceptable salt, for example,an acid-addition salt of a compound of the Formula I, with an inorganicor organic acid such as hydrochloric, hydrobromic, sulfuric,trifluoroacetic, citric or maleic acid. Nowhere in WO2005/028469 isthere a suggestion of a salt with fumaric acid. Compound (I) isdisclosed in Example 1 of WO2005/028469 and is isolated as the freebase. There is no disclosure in WO2005/028469 of any specific salt ofCompound (I).

We have found that Compound (I) is crystalline with some amorphouscharacter as shown in the XRPD of FIG. 1. Differential scanningcalorimetry (FIG. 2A) on Compound (I) shows a broad endotherm with anonset of 76.2° C., which is likely to be due to solvent loss, mostlikely water, followed by a melting endotherm with an onset of 126.2° C.Thermogravimetric analysis on Compound (I) (FIG. 2B) shows a weight lossis of 1.2% between 25° C. and 95° C.

Dynamic Vapour Sorption (FIG. 3) shows moisture uptake of approximately1.9% w/w at 80% relative humidity, accordingly Compound (I) ismoderately hygroscopic.

We have found that Compound (I) has a relatively low intrinsicdissolution rate, particularly at pH below 6.0 and has a high cellularpermeability. The low solubility and high permeability suggest a BCSclassification of Class II for Compound (I). Therefore, the dissolutioncharacteristics of the compound may be critical in controlling drugabsorption and inter patient variability, especially at higher doses.These findings together with the facts that Compound (I) is partiallyamorphous and is hygroscopic has resulted in the need to findalternative forms of Compound (I) with improved properties.

We have surprisingly found that the difumarate salt of Compound (I) hasfavourable properties compared to Compound (I). Compound (I) difumaratehas a favourable dissolution profile exhibiting, high aqueous solubilityand a good intrinsic dissolution rate. Furthermore, the Compound (I)difumarate exhibits favourable solid-state properties, for example highcrystallinity, low hygroscopicity and/or favourable thermal properties,such as a high melting point.

SUMMARY

The present invention relates to a salt of4-(3-chloro-2-fluoroanilino)-7-methoxy-6-{[1-(N-methylcarbamoylmethyl)piperidin-4-yl]oxy}quinazolinehereafter “Compound (I)”, more particularly to the difumarate salt ofCompound (I). The salt is expected to be useful for the treatment orprophylaxis of conditions mediated alone or in part by erbB receptorsignalling, particularly proliferative diseases such as cancer. Theinvention also relates to a pharmaceutical composition comprising thesalt and to the use thereof in the manufacture s of a medicament for usein the treatment or prophylaxis of cancer, such as breast cancer.

BRIEF DESCIPTION OF THE FIGURES

FIG. 1 shows an X-ray powder diffraction pattern (XRPD) for Compound (I)free form. The x-axis shows the 2-theta value and the y-axis the counts.

FIG. 2A shows a differential scanning calorimetry trace on Compound (I)free form. The x-axis shows the temperature and time, the y-axis showspower in mW. The text on the figure shows the onset temperature of theendotherms and the integral (mJ) of the curves.

FIG. 2B is a thermogravimetric trace for Compound (I) free form. Thex-axis shows temperature and time, the y-axis shows weight in mg. Thetext in this graph shows is the % weight loss and absolute weight lossfrom the sample for the event between about 30 and 80° C.

FIG. 3 shows a dynamic vapour sorption isotherm plot for Compound (I)free form. The x-axis shows the % relative humidity, the y-axis showsthe % change in mass of the sample. “sorp” refers to an adsorption cycleand “desorp” to a desorption cycle.

FIG. 4 shows an X-ray powder diffraction pattern (XRPD) for Compound (I)difumarate Form A. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 5 shows a differential scanning calorimetry trace on Compound (I)difumarate Form A. The x-axis shows the temperature and time, the y-axisshows power in mW. The text on the figure shows the onset temperature ofthe melting endotherm and the integral (mJ) of the curve.

FIG. 6 shows a thermogravimetric trace for Compound (I) difumarate FormA. The x-axis shows temperature and time, the y-axis shows weight in mg.The text in this graph shows the % weight loss and absolute weight lossfrom the sample between about 30 and 80° C.

FIG. 7 shows a dynamic vapour sorption isotherm plot for Compound (I)difumarate Form A. The x-axis shows the % relative humidity, the y-axisshows the % change in mass of the sample. “sorp” refers to an adsorptioncycle and “desorp” to a desorption cycle.

FIG. 8 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form B. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 9 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form C. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 10 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form D. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 11 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form E. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 12 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form F. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 13 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form G. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 14 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form H. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 15 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form I. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 16 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form J. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 17 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form K. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 18 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form L. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 19 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form M. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 20 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form N. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 21 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form O. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 22 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form P. The x-axis shows the 2-theta value and the y-axis thecounts.

FIG. 23 shows an X-ray powder diffraction pattern for Compound (I)difumarate Form Q. The x-axis shows the 2-theta value and the y-axis thecounts.

DETAILED DESCRIPTION

Accordingly a first aspect of the present invention provides Compound(I) difumarate.

Suitably the Compound (I) difumarate is crystalline. Therefore accordingto a further aspect of the present invention there is providedcrystalline Compound (I) difumarate.

The Compound (I) difumarate may exist in solvated as well as unsolvatedforms such as, for example, hydrated forms. It is to be understood thatthe invention encompasses all such solvated and unsolvated forms ofCompound (I) difumarate.

We have found that a particular crystalline form of Compound (I)difumarate, hereafter “Form A” is characterised in that it provides anX-ray powder diffraction pattern substantially as shown in FIG. 4. Themost prominent peaks of Form A are shown in Table 1.

TABLE 1 The most prominent X-Ray Powder Diffraction peaks for Form AAngle Intensity 2-Theta° (2θ) Count 6.3 W 7.1 S 8.7 S 10.2 M 10.6 M 11.9S 12.4 S 12.6 M 13.0 S 13.4 M 14.1 S 14.4 M 14.9 S 15.4 M 16.3 M 16.9 M17.3 M 18.8 S 19.2 M 20.3 VS 20.5 VS 21.2 S 21.5 S 21.9 VS 22.7 M 23.0 S23.5 S 24.0 VS 24.7 S 25.5 S 26.1 M 26.4 VS 27.4 S 28.2 M 29.5 S 29.9 S30.8 M 31.5 M 31.8 M 32.2 M 32.7 M 33.0 M 33.5 M 34.0 M 34.7 M 35.1 M35.5 M 35.8 M 36.6 M 37.3 M 38.3 M 39.2 M 39.7 M

In Table 1 the following abbreviations are used: VS=very strong;S=strong; M=medium and W=weak.

According to a further aspect of the invention there is provided Form A,wherein said Form A has an X-ray powder diffraction pattern with atleast one specific peak at about 2-theta=26.4°.

According to a further aspect of the invention there is provided Form A,wherein said

Form A Agent has an X-ray powder diffraction pattern with at least onespecific peak at about 2-theta=26.4°, 14.9° or 7.1°.

According to a further aspect of the invention there is provided Form A,wherein said Form A has an X-ray powder diffraction pattern withspecific peaks at about 2-theta=26.4°, 14.9° and 7.1°.

According to a further aspect of the invention there is provided Form A,wherein said Form A has an X-ray powder diffraction pattern with atleast one specific peak at about 2-theta=26.4°, 24.0°, 14.9°, 12.4° or7.1°.

According to a further aspect of the invention there is provided Form A,wherein said Form A has an X-ray powder diffraction pattern withspecific peaks at about 2-theta=26.4°, 24.0°, 14.9°, 12.4° and 7.1°.

According to a further aspect of the invention there is provided Form A,wherein said Form A has an X-ray powder diffraction pattern with atleast one specific peak at about 2-theta=26.4°, 24.0°, 23.0°, 21.2°,17.3°, 15.4°, 14.9°, 13.0, 12.4° or 7.1°.

According to a further aspect of the invention there is provided Form A,wherein said Form A has an X-ray powder diffraction pattern withspecific peaks at about 2-theta=is 26.4°, 24.0°, 23.0°, 21.2°, 17.3°,15.4°, 14.9°, 13.0°, 12.4° and 7.1°.

According to another aspect of the invention there is provided Form A,wherein said Form A has an X-ray powder diffraction patternsubstantially the same as the X-ray powder diffraction pattern shown inFIG. 4.

Suitably Form A is substantially free of other forms of Compound (I)difumarate. For example, at least 80% of the Compound (I) difumarate isin the form of Form A, particularly at least 90%, more particularly, atleast 95% and still more particularly at least 99% of the Compound (I)difumarate is in the form of Form A. In a particular embodiment at least98% of the Compound (I) difumarate is in the form of Form A. Referenceherein to, for example, 80% of the Compound (I) difumarate being in theform of Form A, refer to the % by weight of the Compound (I) difumarate.

The DSC thermogram for Form A is shown in FIG. 5 hereinafter. Form A,shows a sharp melting endotherm with an onset temperature of about210.4° C., as determined by differential scanning calorimetry (DSC)analysis using a Mettler DSC820e apparatus as described in the Examples.Accordingly Form A has a melting point of about 210° C.

We have found that Compound (I) difumarate may exist in othercrystalline forms, for example the Forms B to Q described in theExamples herein. According to a further aspect of the invention there isprovided crystalline Compound (I) difumarate selected from any one ofForm B to Form Q described herein. Suitably each of the crystallineCompound (I) difumarate forms described is substantially free of otherforms of Compound (I) difumarate. For example, at least 80% of theCompound (I) difumarate is in the desired form, particularly at least90%, more particularly, at least 95% and still more particularly atleast 99% of the Compound (I) difumarate is in the desired crystallineform s of the difumarate.

The crystalline forms of Compound (I) difumarate described herein arecrystalline. Suitably the degree of crystallinity as determined by X-raypowder diffraction data is for example greater than about 60%, such asgreater than about 80%, particularly greater than about 90% and moreparticularly greater than about 95%. In embodiments of the invention,the degree of crystallinity as determined by X-ray powder diffractiondata is greater than about 98%, wherein the % crystallinity refers tothe % by weight of the total sample mass which is crystalline.

In the preceding paragraphs defining the X-ray powder diffraction peaksfor the crystalline forms of Compound (I), the term “at about” is usedin the expression “ . . . at is about 2-theta= . . . ” to indicate thatthe precise position of peaks (i.e. the recited 2-theta angle values)should not be construed as being absolute values because, as will beappreciated by those skilled in the art, the precise position of thepeaks may vary slightly between one measurement apparatus and another,from one sample to another, or as a result of slight variations inmeasurement conditions utilised. It is also stated in the precedingparagraphs that the Compound (I) difumarate Form A provides X-ray powderdiffraction patterns ‘substantially’ the same as the X-ray powderdiffraction patterns shown in FIG. 4, and has substantially the mostprominent peaks (2-theta angle values) shown in Table 1. It is to beunderstood that the use of the term ‘substantially’ in this context isalso intended to indicate that the 2-theta angle values of the X-raypowder diffraction patterns may vary slightly from one apparatus toanother, from one sample to another, or as a result of slight variationsin measurement conditions utilised, so the peak positions shown in theFigures or quoted in the Tables are again not to be construed asabsolute values.

In this regard, it is known in the art that an X-ray powder diffractionpattern may be obtained which has one or more measurement errorsdepending on measurement conditions (such as equipment or machine used).In particular, it is generally known that intensities in an X-ray powderdiffraction pattern may fluctuate depending on measurement conditionsand sample preparation. For example, persons skilled in the art of X-raypowder diffraction will realise that the relative intensity of peaks canbe affected by, for example, grains above 30 microns in size andnon-unitary aspect ratios, which may affect analysis of samples. Theskilled person will also realise that the position of reflections can beaffected by the precise height at which the sample sits in thediffractometer and the zero calibration of the diffractometer. Thesurface planarity of the sample may also have a small effect. Hence aperson skilled in the art will appreciate that the diffraction patterndata presented herein is not to be construed as absolute (for furtherinformation see Jenkins, R & Snyder, R. L. ‘Introduction to X-Ray PowderDiffractometry’ John Wiley & Sons, 1996). Therefore, it shall beunderstood that the crystalline forms of Compound (I) difumaratedescribed herein are not limited to the crystals that provide X-raypowder diffraction patterns identical to the X-ray powder diffractionpattern shown in FIG. 4, and any crystals providing X-ray powderdiffraction patterns substantially the same as those shown in FIG. 4fall within the scope of the present invention. A person skilled in theart of X-ray powder diffraction is able to judge the substantialidentity of X-ray powder diffraction patterns.

Generally, a measurement error of a diffraction angle in an X-ray powderdiffractogram is about 2-theta=0.5° or less, and such degree of ameasurement error should be taken into account when considering theX-ray powder diffraction patterns in FIGS. 1 and 4, and wheninterpreting the peak positions referred to in the text above and inTable 1.

The melting points and DSC data described herein were determined using aMettler

DSC820e apparatus, the use of which is described in more detailhereinafter. A person skilled in the art will appreciate that slightvariations in the melting point measured by DSC may occur as a result ofvariations in sample purity, sample preparation and the measurementconditions (e.g. heating rate). It will be appreciated that alternativereadings of melting point may be given by other types of equipment or byusing conditions different to those described hereinafter. Hence themelting point and endotherm figures quoted herein are not to be taken asabsolute values and such measurement errors are to be taken into accountwhen interpreting DSC data. Typically, melting points may vary by ±0.5°C. or less.

The crystalline forms of Compound (I) difumarate, such as Form Aaccording to the invention may also be characterised and/ordistinguished from other physical forms using other suitable analyticaltechniques, for example NIR spectroscopy or solid state nuclear magneticresonance spectroscopy.

The chemical structure of Compound (I) difumarate of the presentinvention can be confirmed by routine methods for example proton nuclearmagnetic resonance (NMR) analysis.

Synthesis of Compound (I)

Compound (I) may be synthesised using the methods described inWO2005/028469 or as illustrated in the Examples herein.

WO2005/028469 discloses as Example 1 therein the preparation of Compound(I) as follows:

2-Chloro-N-methylacetamide (32 mg, 0.3 mmol) was added to a mixture of4-(3-chloro-2-fluoroanilino)-7-methoxy-6-[(piperidin-4-yl)oxy]quinazoline(120 mg, 0.3 mmol), potassium iodide (16 mg, 0.1 mmol), and potassiumcarbonate (50 mg, 0.36 mmol) in acetonitrile (5 ml). The mixture washeated at reflux for one hour. After evaporation of the solvents undervacuum, the residue was taken up in dichloromethane. The organicsolution was washed with water and brine, dried over magnesium sulfate.After is evaporation of the solvents under vacuum, the residue waspurified by chromatography on silica gel (eluant: 1% to 2% 7N methanolicammonia in dichloromethane) to give Compound (I).

We have found that reaction of 2-chloro-N-methylacetamide directly with4-(3-chloro-2-fluoroanilino)-7-methoxy-6-[(piperidin-4-yl)oxy]quinazolineavoids the use of potassium iodide. Furthermore, crystallisation ofCompound (I) from certain solvents provides Compound (I) in high purity.The new process is therefore expected to be suitable for large-scalemanufacture of Compound (I).

Accordingly as a further aspect of the present invention there isprovided a method for preparing Compound (I) comprising:

(i) reacting 2-chloro-N-methylacetamide with4-(3-chloro-2-fluoroanilino)-7-methoxy-6-[(piperidin-4-yl)oxy]quinazolinedihydrochloride in the presence of a suitable base;

(ii) adding a solvent selected from ethanol, water and methanol, or amixture thereof to the reaction mixture from step (i) to effectcrystallisation of Compound (I); and

(iii) isolating the Compound (I).

The reaction in Step (i) is conveniently carried out is a suitable inertsolvent such as those described in Process (c) on page 30 ofWO2005/028469. For example, the reaction may be carried out usingacetonitrile as the solvent. The reaction is carried out in the presenceof a suitable base, for example one of the bases described in Process(c) on page 30 of WO2005/028469, such as triethylamine. Suitably thereaction is carried out at an elevated temperature, for example at about75° C.

In one embodiment in Step (ii) of the process the solvent is water.Suitably in this embodiment when Step (i) is carried out in acetonitrilethe volume ratio of water:acetonitrile is approximately 1:3.

In another embodiment of the invention in Step (ii) of the process thesolvent is ethanol. Suitably in this embodiment when Step (i) is carriedout in acetonitrile the volume ratio of ethanol:acetonitrile isapproximately 3.5:7.

In a further embodiment of the invention in Step (ii) of the process thesolvent is a mixture of ethanol and water. Suitably in this embodimentthe volume ratio of ethanol to water is about 20:1 to about 30:1, forexample about 21.9:1 to 25:1. When Step (i) is carried out inacetonitrile suitably approximately 3.5 volumes of ethanol and 0.15volumes of water are added to 7 volumes of acetonitrile to effectcrystallisation.

As will be understood reference to a volume ratio of for example ofwater:acetonitrile being 1:3 means that 1 volume of water is added to 3volumes of the acetonitrile present in the reaction vessel following thecompletion of Step (i) of the process.

In one embodiment, in Step (ii) of the process the reaction mixture fromStep (i) is cooled to about 70° C. and ethanol is added. The reactionmixture is then cooled to about 45° C. and the water is added to effectcrystallisation of the Compound (I). If required the reaction mixturemay be seeded with Compound (I) to help initiate the crystallisation.The reaction mixture is then cooled to about 20° C. to complete thecrystallisation.

The isolation of the Compound (I) in Step (iii) may be carried out usingconventional methods, for example filtration and drying of the Compound(I).

The4-(3-chloro-2-fluoroanilino)-7-methoxy-6-[(piperidin-4-yl)oxy]quinazolinedihydrochloride used as the starting material may be prepared asdescribed in the Examples herein. For example, by adding hydrochloricacid to6-{[(1-tert-butoxycarbonyl)piperidin-4-yl]oxy}-4-(3-chloro-2-fluoroanilino)-7-methoxyquinazoline.Conveniently the reaction is carried out in a suitable solvent, forexample ethanol, acetonitrile, tetrahydrofuran, 2-methyltetrahydrofuran,n-propanol, methanol, 1-butanol, ethyl acetate, tent-butyl acetate,isopropanol or industrial methylated spirit. A particular solvent isethanol or more particularly industrial methylated spirit.

The6-{[(1-tert-butoxycarbonyl)piperidin-4-yl]oxy}-4-(3-chloro-2-fluoroanilino)-7-methoxyquinazolinemay be prepared using the method described in Example 1 of WO2005/028469by reacting 4-(3-Chloro-2-fluoroanilino)-6-hydroxy-7-methoxyquinazolinewith tert-Butyl (4-methanesulfonyloxy)piperidine-1-carboxylate, whereinthe reaction is carried out in the presence of cesium fluoride using DMAas a solvent and at a temperature of 85° C.

This reaction may also be carried out in the presence ofN-methylpyrrolidone (NMP) in the presence of a suitable base such aspotassium carbonate. This reaction is suitably performed at an elevatedtemperature as described in the Examples herein.

However, we have found that by carrying the reaction out in the presenceof certain solvents provides the product in good form. Furthermore someof these solvents are expected to be suitable for large-scalemanufacture of the product.

Accordingly, as a further aspect of the present invention there isprovided a process for the preparation of6-{[(1-tert-butoxycarbonyl)piperidin-4-yl]oxy}-4-(3-chloro-2-fluoroanilino)-7-methoxyquinazolinecomprising:

(i) reacting 4-(3-chloro-2-fluoroanilino)-6-hydroxy-7-methoxyquinazolinewith tert-butyl (4-methanesulfonyloxy)piperidine-l-carboxylate in thepresence of a suitable base, wherein the reaction is carried out in asolvent selected from N-methylpyrrolidone or an alcohol selected frommethanol, ethanol, isopropyl alcohol, n-propanol and industrialmethylated spirit; and

(ii) crystallising the6-{[(1-tert-butoxycarbonyl)piperidin-4-yl]oxy}-4-(3-chloro-2-fluoroanilino)-7-methoxyquinazoline.

Step (i) of this process is carried out in the presence of a suitablebase, for example those described in WO2005/028469, such as potassiumcarbonate. The reaction is suitably carried out at elevated temperatureconveniently at the reflux temperature.

If required water can be added to the solvents used in Step (i) to aidprocessing, for example to increase the mobility of the reactionmixture. In one embodiment Step (i) of the reaction is carried out in analcohol selected from methanol, ethanol, isopropyl alcohol, n-propanoland industrial methylated spirit, optionally in the presence of water.In a further embodiment the reaction Step (i) is carried out in amixture of ethanol and water. When a mixture of ethanol and water isused, the volume ratio of ethanol to water in Step (i) is not critical,for example a volume ratio of ethanol to water may be up to about 10:2is suitable, such as about 10:1.

The crystallisation in Step (ii) of the process is conveniently carriedout by cooling the reaction mixture from Step (i) (for example coolingto about 70° C.) and adding water to the mixture to effectcrystallisation. The product may then be isolated by conventionalmethods such as those described in the Examples.

Compound (I) may also be prepared according to the process illustratedin Reaction Scheme 1:

Notes on Reaction Scheme 1:

Step (i): Lg¹ is a suitable leaving group, for example, a halogeno,alkanesulfonyloxy or arylsulfonyloxy group, for example a chloro, bromo,methanesulfonyloxy, 4-nitrobenzenesulfonyloxy or toluene-4-sulfonyloxygroup (suitably a methanesulfonyloxy, 4-nitrobenzenesulfonyloxy ortoluene-4-sulfonyloxy group, for example Lg¹ is methanesulfonyloxy).

Pg¹ is a suitable amine protecting group. Such groups are well known,for example as described in one of the many general texts on thesubject, such as, ‘Protective Groups in Organic Synthesis’ by TheodoraGreen (publisher: John Wiley & Sons). Examples of amino protectinggroups include an acyl group, for example an alkanoyl group such asacetyl, an alkoxycarbonyl group, for example a methoxycarbonyl,ethoxycarbonyl or tert-butoxycarbonyl group, an arylmethoxycarbonylgroup, for example benzyloxycarbonyl, or an aroyl group, for examplebenzoyl. A particular example of Pg¹ is tert-butoxycarbonyl.

The reaction is suitably carried out in the presence of a base, forexample a is carbonate such as potassium carbonate. The reaction isconveniently carried out in the presence of a suitable inert solvent,for example an alcohol such as isopropanol. The reaction is suitablycarried out at elevated temperature, conveniently at the refluxtemperature of the solvent.

Step (ii): The protecting group Pg¹ is removed using conventionalmethods. For example when Pg¹ is tert-butoxycarbonyl it may be removedby treatment with a suitable acid as hydrochloric, sulfuric orphosphoric acid or trifluoroacetic acid.

Step (iii): Lg² is a suitable leaving group, for example, a halogeno,such as chloro. The reaction is suitably carried out in the presence ofa suitable base such as a carbonate, an organic amine or an alkoxide.Suitable bases include potassium carbonate or triethanolamine. Thereaction is conveniently carried out in the presence of an inert solventsuch as acetonitrile or an alcohol such as ethanol. The reaction issuitably performed at an elevated temperature, conveniently the refluxtemperature of the solvent.

Step (iv): Nitration may be effected using well known methods for thenitration of aromatic rings, for example by treating the2-[4-(5-Cyano-2-methoxyphenoxy)piperidin-1-yl]-N-methylacetamide (4)with nitric acid in the presence of sulfuric acid using well knownconditions for such reactions and as illustrated in the Examples.

Step (v): Reduction reactions suitable for reducing nitro groups toamines are well known, for example by reduction in the presence of asuitable reducing agent such as sodium dithionite. This reaction issuitably carried out in the presence of an aqueous solvent, for exampleaqueous methanol. The reaction is conveniently performed at elevatedtemperature for example 40 to 60° C. Alternatively, reduction may beeffected by hydrogenation, for example by catalytic hydrogenation withusing a suitable catalyst such as a palladium on carbon catalyst, forexample a 10% palladium on carbon catalyst. The hydrogenation isconveniently carried out in a suitable solvent such as methanol.

Step (vi):2-[4-(4-Amino-5-cyano-2-methoxyphenoxy)piperidin-1-yl]-N-methylacetamide(6) is reacted with N,N-dimethylformamide dimethyl acetal. The reactionis conveniently carried out in the presence of a suitable solvent suchas an ether, for example, 2-methyltetrahydrofuran or an aromatichydrocarbon such as toluene. The reaction is suitably performed at anelevated temperature, for example at about 70 to 105° C., suitably about76° C.

Step (vii) The reaction is suitably carried out in the presence of asuitable acid, such as one or more acids selected from acetic,propanoic, succinic, fumaric and citric acid. In is one example the acidis acetic acid. The reaction is suitably carried out in the presence ofan inert solvent, for example an aromatic hydrocarbon solvent such asmethoxybenzene. The reaction is suitably carried out at elevatedtemperature, for example from about 90 to about 120° C., suitably atabout 90° C.

The process described in Reaction Scheme 1 forms a further aspect of thepresent invention. Accordingly, there is provided a method for preparingCompound (I) comprising reacting2-[4-(5-cyano-4-{[(dimethylamino)methylene]amino}-2-methoxyphenoxy)piperidin-1-yl]-N-methylacetamide(7) with 3-chloro-2-fluoroaniline in the presence of a suitable acid.

Suitable reaction conditions are as hereinbefore described in relationto Step (vii) of Reaction Scheme 1.

Certain intermediates shown in Reaction Scheme 1 are novel and form afurther aspect of the present invention. Accordingly another aspect ofthe invention provides a compound selected from any one of compounds 2,3, 4, 5, 6 and 7 in Reaction Scheme 1, or a salt thereof; wherein Pg¹ isas hereinbefore defined (for example tert-butoxycarbonyl). Some of theintermediates such as compound (7) in Reaction Scheme 1 may havegeometric isomeric centres (E- and Z-isomers). It is to be understoodthat the present invention encompasses all such geometric isomers andmixtures thereof.

Synthesis of Compound (I) Difumarate Form A

According to a further aspect of the present invention there is provideda process for the preparation of Compound (I) difumarate (Form A)comprising:

(i) reacting Compound (I) with a sufficient quantity of fumaric acid toform the s difumarate salt;

(ii) crystallising the Form A; and

(iii) isolating the Form A.

Notes on Step (i)

Conveniently the reaction with the fumaric acid is carried out in asuitable solvent, for example, selected from methanol, ethanol,1-butanol, 2-butanol and diacetone alcohol. The reaction may also becarried out in a mixture of suitable solvents, for example a mixtureselected from methyl ethyl ketone and dimethylformamide; methyl ethylketone and tetrahydrofuran; methyl ethyl ketone and methanol; methylethyl ketone and isopropanol; ethanol and dimethyl sulfoxide; ethanoland tetrahydrofuran; ethanol and isopropanol; 1-butanol anddimethylformamide; 1-butanol and dimethyl sulfoxide; 1-butanol andtetrahydrofuran; 1-butanol and methanol; 1-butanol and isopropanol;ethylacetate and dimethylformamide; ethyl acetate and methanol; ethylacetate and isopropanol; and methanol and isopropanol.

In one embodiment the reaction in Step (i) is carried out in water.

In one embodiment the reaction in Step (i) is carried out in a mixtureof solvents comprising methyl ethyl ketone and a solvent selected fromdimethylformamide, tetrahydrofuran, methanol and isopropanol.

In another embodiment the reaction in Step (i) is carried out in amixture of solvents comprising ethanol and a solvent selected fromdimethyl sulfoxide, tetrahydrofuran and isopropanol.

In another embodiment the reaction in Step (i) is carried out in amixture of solvents comprising 1-butanol and a solvent selected fromdimethylformamide, dimethyl sulfoxide, tetrahydrofuran, methanol andisopropanol.

In another embodiment the reaction in Step (i) is carried out in amixture of solvents comprising ethyl acetate and a solvent selected fromdimethyl formamide, methanol and isopropanol.

In another embodiment the reaction in Step (i) is carried out in amixture of solvents comprising ethyl acetate and isopropanol.

In another embodiment the reaction in Step (i) is carried out in amixture of solvents comprising methanol and isopropanol.

In those embodiments where Step (i) of the reaction is carried out in amixture of solvents comprising methanol and isopropanol, the volumeratio of isopropanol to methanol is suitably in the range of about 3.4:1to about 1.0:1, for example about 1.5:1 to about 1.0:1. This reaction issuitably carried out at an elevated temperature, for example at atemperature in excess of 60° C., suitably from 65° C. to the refluxtemperature of the solvent. Conveniently the Compound (I) is dissolvedor dispersed in the isopropanol and this mixture is reacted with asolution or dispersion of the fumaric acid in the methanol.

In those embodiments where Step (i) of the reaction is carried out in amixture of solvents comprising ethyl acetate and isopropanol, the volumeratio of ethyl acetate to isopropanol is suitably in the range of about5.1:1 to 1.9:1, for example about 3.9:1 to 1.9:1 such as about 2.1:1.This reaction is suitably carried out at a temperature of about 20 toabout 73° C., for example at about 40° C. Conveniently the Compound (I)is dissolved or dispersed in the ethyl acetate and this mixture isreacted with a solution or dispersion of the fumaric acid in theisopropanol. Alternatively, Compound (I) may be pre-dissolved in amixture of ethyl acetate and isopropanol. If required, the isopropanolmay by removed following dissolution of the Compound (I) usingconventional methods such as distillation.

When preparing the solution or dispersion of fumaric acid in an alcoholsuch as methanol or isopropyl alcohol, it may be necessary to heat themixture to effect dissolution of the fumaric acid. However, excessiveheating and/or holding the mixture at elevated temperature for prolongedperiods of time should be avoided to minimise ester formation.

Generally in Step (i) of the process Compound (I) is reacted with atleast 2 molar equivalents of fumaric acid, for example from about 2 toabout 3, particularly about 2 to about 2.7 molar equivalents of fumaricacid. However, lower quantities of fumaric acid can be used in certainsolvent systems. For example, when the reaction is carried out in amixture of ethyl acetate and isopropanol we have found that Compound (I)difumarate can be prepared when the molar ratio of fumaric acid toCompound (I) is greater than or equal to 1.725.

Notes on Step (ii)

Crystallisation of the Form A may be effected using known methods forcrystallisation of a compound from solution. For example by causingsupersaturation of the solution containing the salt. Supersaturation maybe achieved by, for example, cooling the solution, evaporating solventfrom the solution or by addition of a suitable anti-solvent to thesolution.

In one embodiment crystallisation is effected by cooling the solution.For example, when Step (i) of the reaction is carried out in a mixtureof methanol and isopropyl alcohol, the reaction mixture is cooled toabout 30° C. over a period of about 90 minutes and is held at 30° C. forabout 30 minutes. The reaction mixture may then be further cooled over aperiod of about 2 hours to a temperature of about 0° C. and is held atthe temperature for a sufficient time to allow completion ofcrystallisation, for example about 1 hour.

Alternatively, when Step (i) of the reaction is carried out in a mixtureof ethyl acetate and isopropyl alcohol, the reaction mixture is cooledto about 20° C. (for example is from about 40° C. to about 20° C. over aperiod of about 1 hour). The reaction mixture is then held at 20° C. fora sufficient time to effect crystallisation. Suitably, the reactionmixture is held at about 20° C. for at least 10 hours, for example forabout 13.5 hours.

In another embodiment when Step (i) of the reaction is carried out in amixture of methanol and isopropyl alcohol, crystallisation may beeffected by removing a proportion of the solvent to causesupersaturation of the remaining reaction mixture. The solvent may beremoved by evaporation or distillation. Suitably about 55 to 65% byweight of the solvent is removed, for example about 62%. If requiredfurther isopropanol may be added to the mixture followed by distillationof approximately the same weight of solvent. For example approximately50 to 60% of additional isopropanol may be added to the mixture, whereinthe % is the % by weight of the solvent remaining in the reaction vesselfollowing the first distillation. Following addition of the isopropanola similar weight of solvent is removed by distillation. Crystallisationmay be completed by adding further isopropanol and cooling the mixtureto about 0° C. over a period of about 8 hours.

Generally the Form A will self crystallise in Step (ii) of the process,but as will be appreciated by a person skilled in the art, seeding withForm A may be used in order to promote crystallisation. If required,seed crystals could be prepared using the method described above andillustrated in the Examples for the preparation of Compound (I)difumarate Form A.

Notes on Step (iii)

Any suitable method known in the art for isolating crystalline materialsfrom a solution may be used in Step (iii) of the process. Suitably, theForm A is collected by filtration. Following isolation of the Form A,the salt may be washed with a suitable solvent, for example coldisopropanol. Following isolation the Form A may be dried usingconventional methods, for example vacuum drying.

Accordingly in one embodiment of the invention there is provided aprocess for the preparation of Compound (I) difumarate (Form A)comprising:

(i) reacting a solution or suspension of Compound (I) in isopropanolwith at least 2 molar equivalents fumaric acid in methanol,

-   -   wherein the volume ratio of isopropanol to methanol is from        3.4:1 to about 1.0:1, for example about 1.5:1 to about 1.0:1,    -   and wherein the reaction is carried out at a temperature of at        least 60° C.;

(ii) crystallising the Form A; and

(iii) isolating the Form A.

Suitable conditions for crystallisation and isolation of the Form A areas hereinbefore defined.

Accordingly in another embodiment of the invention there is provided aprocess for the preparation of Compound (I) difumarate (Form A)comprising:

(i) reacting a solution or suspension of Compound (I) in ethyl acetatewith at least a 1.725 molar equivalents fumaric acid in isopropanol(suitably at least 2 molar equivalents of fumaric acid),

wherein the volume ratio of ethyl acetate to isopropanol is suitablyfrom about 5:1 to 1:1, for example about 5.1:1 to 1.9:1 such as about2.1:1 and wherein the reaction is carried out at a temperature of about20 to about 73° C. (for example about 40° C.);

(ii) cooling the reaction mixture from Step (i) to about 20° C. andholding the mixture at this temperature to effect crystallisation of theForm A; and

(iii) isolating the Compound (I) difumarate Form A.

Suitable conditions for isolation of the Form A are as hereinbeforedefined.

In another embodiment of the invention there is provided a process forthe preparation of Compound (I) difumarate (Form A) comprising:

(i) reacting Compound (I) in water with at least 2 molar equivalents offumaric acid (for example at least 2.05, such as about 2.1 molarequivalents of fumaric acid), and wherein the reaction is carried out atabout 85° C.;

(ii) cooling the reaction mixture from Step (i) to about 60° C.; and

(iii) isolating the Compound (I) difumarate Form A.

Suitably in Step (ii) the reaction mixture is cooled slowly to about 60°C., for example at a cooling rate of about 1° C/minute. If requiredcrystallisation of the Form A can be induced by adding seed crystals ofthe Form A during cooling of the mixture. Suitably the Form A seedcrystals are added when the reaction mixture has been cooled to about77° C. Suitable conditions for isolation of the Form A in Step (iii) areas hereinbefore described.

Crystalline Compound (I) difumarate Forms B to P may be prepared by, forexample, the methods described herein in the Examples.

Pharmaceutical Compositions

According to a further aspect of the invention there is provided apharmaceutical composition which comprises Compound (I) difumarate inassociation with a pharmaceutically-acceptable diluent or carrier. TheCompound (I) difumarate may be used in the composition in any of theforms described herein, for example Form A.

The compositions of the invention may be in a form suitable for oral use(for example as tablets, lozenges, hard or soft capsules, aqueous oroily suspensions, emulsions, dispersible powders or granules, syrups orelixirs), for topical use (for example as creams, ointments, gels, oraqueous or oily solutions or suspensions), for administration byinhalation (for example as a finely divided powder or a liquid aerosol),for administration by insufflation (for example as a finely dividedpowder) or for parenteral administration (for example as a sterileaqueous or oily solution for intravenous, subcutaneous, intramuscular orintramuscular dosing or as a suppository for rectal dosing).

The compositions of the invention may be obtained by conventionalprocedures using conventional pharmaceutical excipients, well known inthe art. Thus, compositions intended for oral use may contain, forexample, one or more colouring, sweetening, flavouring and/orpreservative agents.

For example Compound (I) difumarate is suitably formulated as a tabletusing the following excipients:

Tablet Core:

Compound (I) difumarate (for example Form A);

lactose;

microcrystalline cellulose;

crospovidone;

polyvidone (PVP); and

magnesium stearate

The tablet core may be coated with a film-coating, such as an HPMC basedfilm coating, which coating optionally contains one or more colorantsand/or light protective agents.

The tablets may be prepared using conventional methods and asillustrated in the Examples. If required the Compound (I) difumarate maybe milled prior to formulation into the tablet to provide a uniformparticle size distribution of the Compound (I) difumarate in the tablet.For example the Compound (I) difumarate may be milled to is provide anaverage particle size of about 5 μm. Suitable milling methods are wellknown.

The amount of active ingredient that is combined with one or moreexcipients to produce a single dosage form will necessarily varydepending upon the host treated and the particular route ofadministration. For example, a formulation intended for oraladministration to humans will generally contain, for example, from 0.5mg to 0.5 g of active agent (more suitably from 0.5 to 200 mg, forexample from 1 to 30 mg) compounded with an appropriate and convenientamount of excipients which may vary from about 5 to about 98 percent byweight of the total composition.

The size of the dose for therapeutic or prophylactic purposes ofCompound (I) difumarate will naturally vary according to the nature andseverity of the conditions, the age and sex of the animal or patient andthe route of administration, according to well known principles ofmedicine.

In using Compound (I) difumarate for therapeutic or prophylacticpurposes it will generally be administered so that a daily dose in therange, for example, 0.1 mg/kg to 75 mg/kg body weight is received, givenif required in divided doses. In general lower doses will beadministered when a parenteral route is employed. Thus, for example, forintravenous administration, a dose in the range, for example, 0.1 mg/kgto 30 mg/kg body weight will generally be used. Similarly, foradministration by inhalation, a dose in the range, for example, 0.05mg/kg to 25 mg/kg body weight will be used. Oral administration ishowever preferred, particularly in tablet form. For instance, Compound(I) difumarate could be administered to a warm-blooded animal orally, ata unit dose less than 1 g daily but more than lmg. Particularly Compound(I) difumarate could be administered to a warm-blooded animal, at a unitdose of less than 250 mg per day. In another aspect of the invention,Compound (I) difumarate could be administered to a warm-blooded animal,at a unit dose of less than 160 mg per day. In a further aspect of theinvention, Compound (I) difumarate could be administered to awarm-blooded animal, at a unit dose of less than 50 mg per day. The doseof Compound (I) difumarate may be administered as a single daily dose oras multiple fractions of the total daily dose. For example, the totaldaily dose of io Compound (I) difumarate may be administered as twodoses, which may be the same or different. Suitably however, eachfraction of the total daily dose would be approximately equal. By way ofexample Compound (I) difumarate may be administered as a one or moreoral dosage forms such as a tablet or capsule containing 1.5, 3.7, 14.9,59.6 or 149 mg of Compound (I) difumarate (equivalent to 1, 2.5, 10, 40or 100 mg of Compound (I) is free form). In a further embodiment a doseof Compound (I) difumarate equivalent to 40, 80, 100, 160, 200 or 240 mgof Compound (I) is administered twice a day. In a particular embodimenta dose of Compound (I) difumarate equivalent to 160 mg of Compound (I)is administered twice a day. In a particular embodiment a dose ofCompound (I) difumarate equivalent to 200 mg of Compound (I) isadministered twice a day. In another particular embodiment a dose ofCompound (I) difumarate equivalent to 240 mg of Compound (I) isadministered twice a day.

Biological Assays

The inhibitory activities of Compound (I) and Compound (I) difumaratemay be measured in the assays described in WO2005/028469 or as describedin the Examples herein.

The compounds of the present invention possess anti-proliferativeproperties such as anti-cancer properties that are believed to arisefrom their erbB family receptor tyrosine kinase inhibitory activity, andparticularly a mixed erbB2/ EGF and/or erbB3/EGF profile.

Accordingly, the compounds of the present invention are expected to beuseful in the treatment of diseases or medical conditions mediated aloneor in part by erbB receptor tyrosine kinases, i.e. the compounds may beused to produce an erbB receptor tyrosine kinase inhibitory effect in awarm-blooded animal in need of such treatment. Thus the compounds of thepresent invention provide a method for the treatment of malignant cellscharacterised by inhibition of one or more of the erbB family ofreceptor tyrosine kinases. Particularly the compounds of the inventionmay be used to produce an anti-proliferative and/or pro-apoptotic and/oranti-invasive effect mediated alone or in part by the inhibition of erbBreceptor tyrosine kinases. Particularly, the compounds of the presentinvention are expected to be useful in the prevention or treatment ofthose tumours that are sensitive to inhibition of one or more of theerbB receptor tyrosine kinases, that are involved in the signaltransduction steps which drive proliferation and survival of thesetumour cells. Accordingly the compounds of the present invention areexpected to be useful in the treatment of psoriasis, benign prostatichyperplasia (BPH), atherosclerosis and restenosis and/or cancer byproviding an anti-proliferative effect, particularly in the treatment oferbB receptor tyrosine kinase sensitive cancers. Such benign ormalignant tumours may affect any tissue and include non-solid tumourssuch as leukaemia, multiple myeloma or lymphoma, and also solid tumours,for example bile duct, bone, bladder, brain/CNS, breast, colorectal,endometrial, gastric, head and neck, hepatic, lung, neuronal, isoesophageal, ovarian, pancreatic, prostate, renal, skin, testicular,thyroid, uterine and vulval cancers.

Where cancer is referred to, particularly it refers to oesophagealcancer, myeloma, hepatocellular, pancreatic, cervical cancer, Ewing'stumour, neuroblastoma, Kaposi's sarcoma, ovarian cancer, breast cancer,colorectal cancer, prostate cancer, bladder cancer, melanoma, lungcancer—non small cell lung cancer (NSCLC), and small cell lung cancer(SCLC), gastric cancer, head and neck cancer, brain cancer, renalcancer, lymphoma and leukaemia. In one embodiment it refers to breastcancer, for example hormone receptor-positive breast cancer. In anotherembodiment cancer refers to SCLC, NSCLC, colorectal cancer, ovariancancer and/or breast cancer. In another embodiment cancer refers toSCLC. In another embodiment cancer refers to gastric cancer. Inaddition, it refers to NSCLC. In addition, it refers to colorectalcancer. In addition, it refers to ovarian cancer. In addition, moreparticularly it refers to breast cancer. In addition, more particularlyit refers to hormone receptor positive breast cancer, especially tohormone receptor positive breast cancer in post-menopausal women. In oneembodiment it refers to early stage non-metastatic hormone receptorpositive breast cancer, for example early stage non-metastatic hormonereceptor positive breast cancer in post-menopausal women. Stillfurthermore it refers to early stage non-metastatic estrogen and/orprogesterone receptor positive breast cancer, especially to early stagenon-metastatic estrogen and/or progesterone receptor positive breastcancer in post-menopausal women. In addition, more particularly itrefers to metastatic hormone receptor positive breast cancer, especiallyto metastatic hormone receptor positive breast cancer in post-menopausalwomen. Still furthermore it refers to metastatic estrogen and/orprogesterone receptor positive breast cancer, especially to metastaticestrogen and/or progesterone receptor positive breast cancer inpost-menopausal women. Furthermore, it refers to bladder cancer,oesophageal cancer, gastric cancer, melanoma, cervical cancer and/orrenal cancer. In addition it refers to endometrial, liver, stomach,thyroid, rectal and/or brain cancer. In another embodiment of theinvention, particularly the cancer is in a non-metastatic state. Inanother embodiment of the invention, particularly the cancer is in ametastatic state. In a further embodiment of the invention, particularlythe cancer is in a metastatic state, and more particularly the cancerproduces skin metastases. In a further embodiment of the invention,particularly the cancer is in a metastatic state, and more particularlythe cancer produces lymphatic metastases. In a further embodiment of theinvention, particularly the cancer is in a metastatic state, and moreparticularly the cancer produces brain metastases.

Where the treatment of cancer is referred to particularly this is thetreatment of cancerous tumours expressing one or more of the erbB familyof receptors, for example EFGR, erbB2 and/or erbB3 receptors. Theanti-cancer effect of Compound (I) difumarate according to the inventionmay be measured in terms of one or more of the anti-tumour effect, theextent of the response (for example reduced tumour volume or reducedtumour burden), the response rate, the clinical benefit rate (the sum ofcomplete response, partial response and stable disease) the time todisease progression, progression-free survival and the overall survivalrate. Such clinical trial endpoints are well known and are described infor example the FDA publication “Guidance for Industry Clinical TrialEndpoints for the Approval of Cancer Drugs and Biologics” May 2007(www.fda.gov/CbER/gdlns/clintrialend.htm). The anti-tumour effects ofCompound (I) difumarate according to the invention may be for example,one or more of inhibition of tumour growth, tumour growth delay,regression of tumour, shrinkage of tumour, increased time to regrowth oftumour on cessation of treatment or slowing of disease progression.

The use of Compound (I) difumarate may also have a beneficial effect inpreventing the onset of cancer in warm-blooded animals, such as man.

According to this aspect of the invention there is provided Compound (I)difumarate, for use as a medicament.

According to a further aspect of the invention there is providedCompound (I) difumarate, for use in the production of ananti-proliferative effect in a warm-blooded animal such as man.

Thus according to this aspect of the invention there is provided the useof Compound (I) difumarate in the manufacture of a medicament for use inthe production of an anti-proliferative effect in a warm-blooded animalsuch as man.

According to a further feature of this aspect of the invention there isprovided a method for producing an anti-proliferative effect in awarm-blooded animal, such as man, in need of such treatment whichcomprises administering to said animal an effective amount of Compound(I) difumarate.

According to a further aspect of the invention there is providedCompound (I) difumarate, for use in the prevention or treatment of thosetumours which are sensitive to inhibition of erbB receptor tyrosinekinases, such as a combination of EGFR and erbB2 and/or EGFR and erbB3,that are involved in the signal transduction steps which lead to theproliferation of tumour cells.

According to a further aspect of the invention there is provided the useof Compound (I) difumarate in the manufacture of a medicament for use inthe prevention or treatment of those tumours which are sensitive toinhibition of erbB receptor tyrosine kinases, such as a combination ofEGFR and erbB2 and/or EGFR and erbB3, that are involved in the signaltransduction steps which lead to the proliferation of tumour cells.

According to a further feature of this aspect of the invention there isprovided a method for the prevention or treatment of those tumours whichare sensitive to inhibition of one or more of the erbB family ofreceptor tyrosine kinases, such as a combination of EGFR and erbB2and/or EGFR and erbB3, that are involved in the signal transductionsteps which lead to the proliferation and/or survival of tumour cellswhich comprises administering to said animal an effective amount ofCompound (I) difumarate.

According to a further aspect of the invention there is provided the useof Compound (I) difumarate in the manufacture of a medicament for use inproviding a combined EGFR and erbB2 tyrosine kinase inhibitory effect.

According to a further feature of this aspect of the invention there isprovided a method for providing a combined EGFR and erbB2 tyrosinekinase inhibitory effect which comprises administering to said animal aneffective amount of Compound (I) difumarate.

According to a further feature of this aspect of the invention there isprovided Compound (I) difumarate, for use in providing a combined EGFRand erbB2 tyrosine kinase inhibitory effect.

According to a further aspect of the invention there is provided the useof Compound (I) difumarate in the manufacture of a medicament for use inproviding a tyrosine kinase inhibitory effect on two or more receptorsselected from EGFR, erbB2 and erbB3.

According to a further feature of this aspect of the invention there isprovided a method for providing a tyrosine kinase inhibitory effect ontwo or more receptors selected from EGFR, erbB2 and erbB3, whichcomprises administering to said animal an effective amount of Compound(I) difumarate.

According to a further feature of this aspect of the invention there isprovided Compound (I) difumarate, for use in providing a tyrosine kinaseinhibitory effect on two or more receptors selected from EGFR, erbB2 anderbB3.

According to a further aspect of the invention there is provided the useof Compound (I) difumarate in the manufacture of a medicament for use inthe treatment of a condition (for example a tumour) mediated in whole orpart by the phosphorylation of an erbB2/erbB3 heterodimer.

According to a further feature of this aspect of the invention there isprovided a method for the treatment of a condition (for example atumour) mediated in whole or part by the phosphorylation of anerbB2/erbB3 heterodimer, which comprises administering to said animal aneffective amount of Compound (I) difumarate.

According to a further feature of this aspect of the invention there isprovided Compound (I) difumarate, for use in the treatment of acondition (for example a tumour) mediated in whole or part by thephosphorylation of an erbB2/erbB3 heterodimer.

According to a further aspect of the present invention there is providedthe use of Compound (I) difumarate in the manufacture of a medicamentfor use in the treatment of a cancer (for example a cancer selected fromleukaemia, multiple myeloma, lymphoma, bile duct, bone, bladder,brain/CNS, breast, colorectal, endometrial, gastric, head and neck,hepatic, lung (particularly non-small cell lung cancer), neuronal,oesophageal, ovarian, pancreatic, prostate, renal, skin, testicular,thyroid, uterine and vulval cancer and particularly a cancer selectedfrom breast, gastric, colorectal, head and neck, ovarian and lungcancer, more particularly breast cancer).

According to a further feature of this aspect of the invention there isprovided a method for treating a cancer (for example a cancer selectedfrom leukaemia, multiple myeloma, lymphoma, bile duct, bone, bladder,brain/CNS, breast, colorectal, endometrial, gastric, head and neck,hepatic, lung (particularly non-small cell lung cancer), neuronal,oesophageal, ovarian, pancreatic, prostate, renal, skin, testicular,thyroid, uterine and vulval cancer and particularly a cancer selectedfrom breast, gastric, colorectal, head and neck, ovarian and lungcancer, more particularly breast cancer in a warm-blooded animal, suchas man, in need of such treatment, which comprises administering to saidanimal an effective amount of Compound (I) difumarate.

According to a further aspect of the invention there is providedCompound (I) difumarate, for use in the treatment of a cancer (forexample selected from leukaemia, multiple myeloma, lymphoma, bile duct,bone, bladder, brain/CNS, breast, colorectal, endometrial, gastric, headand neck, hepatic, lung (particularly non-small cell lung cancer),neuronal, oesophageal, ovarian, pancreatic, prostate, renal, skin,testicular, thyroid, uterine and vulval cancer and particularly a cancerselected from breast, gastric, colorectal, head and neck, ovarian andlung cancer, more particularly breast cancer).

As mentioned above the size of the dose required for the therapeutic orprophylactic treatment of a particular disease will necessarily bevaried depending upon, amongst other things, the host treated, the routeof administration and the severity of the illness being treated.

According to a further aspect of the invention there is provided apharmaceutical composition which comprises Compound (I) difumarate, inassociation with a pharmaceutically acceptable diluent or carrier foruse in the treatment of a cancer

For the avoidance of doubt, where the treatment of cancer is indicated,it is to be understood that this also refers to the prevention ofmetastases and the treatment of metastases, i.e. cancer spread.Therefore Compound (I) and Compound (I) difumarate of the presentinvention could be used to treat a patient who has no metastases to stopthem occurring, or to lengthen the time period before they occur, and toa patient who already has metastases to treat the metastases themselves.Furthermore the treatment of cancer also refers to treatment of anestablished primary tumour or tumours and developing primary tumour ortumours. In one aspect of the invention the treatment of cancer relatesto the prevention of metastases. In another aspect of the invention thetreatment of cancer relates to the treatment of metastases. In anotheraspect of the invention the treatment of cancer relates to treatment ofan established primary tumour or tumours or developing primary tumour ortumours. In one embodiment the treatment of cancer relates to anadjuvant treatment. In another embodiment the treatment of cancer refersto the neo-adjuvant treatment of cancer. Accordingly in an embodiment ofthe invention the Compound (I) s difumarate according to the inventionis used as an adjuvant treatment of hormone sensitive breast cancer,particularly as an adjuvant treatment of estrogen receptor positivebreast cancer in post-menopausal women. In another embodiment of theinvention the Compound (I) difumarate according to the invention is usedas a neo-adjuvant treatment of hormone sensitive breast cancer,particularly as a neo-adjuvant treatment of estrogen and/or progesteronereceptor breast cancer in post-menopausal women. In another embodimentthe Compound (I) difumarate is used to treat advanced (metastatic)hormone sensitive (estrogen and/or progesterone receptor positive)breast cancer, particularly advanced estrogen receptor positive cancerin post-menopausal women.

In a further embodiment the Compound (I) difumarate according to theinvention is may be used as a neo-adjuvant therapy in the treatment ofhormone sensitive breast cancer in patients. In another embodiment theCompound (I) difumarate according to the invention is not used as aneo-adjuvant treatment.

The term “adjuvant therapy” refers to a treatment given followingremoval of the primary tumour. Where the cancer is breast cancer,removal of the primary tumour may be effected by, for example, surgery(for example lumpectomy or mastectomy) and/or radiotherapy.

The term “neo-adjuvant therapy” refers to a treatment given prior toremoval of the primary tumour by surgery or radiotherapy.

Herein, the treatment of cancer also refers to the prevention of cancerper se.

In one embodiment of the invention the Compound (I) difumarate is usedin combination with an endocrine agent suitable for use in the treatmentof breast cancer. For example, a combination of Compound (I) difumarateand an endocrine agent selected from an aromatase inhibitor, a selectiveestrogen receptor modulator, an LHRH agonist and an estrogen receptordown-regulator. For example, a combination of Compound (I) difumarateand an aromatase inhibitor. For example a combination of Compound (I)difumarate and tamoxifen. For example a combination of Compound (I)difumarate and anastrozole. For example a combination of Compound (I)difumarate and letrozole. For example a combination of Compound (I)difumarate and exemestane. The combination of Compound (I) difumarateand an endocrine therapy may be particularly suitable for use in thetreatment of breast cancer as described herein. For example, thecombination may be useful in the treatment of metastatic estrogen and/orprogesterone positive breast cancer. Alternatively the combination maybe useful as an adjuvant treatment of breast cancer, particularly as anadjuvant treatment of estrogen and/or progesterone positive breastcancer. The combination may also be useful in the treatment of estrogenand/or progesterone positive breast cancer in patients that have notreceived prior endocrine therapy (for example a selective estrogenreceptor modulator such as tamoxifen, an aromatase inhibitor such asanastrozole or an estrogen receptor down-regulator).

Accordingly, in one embodiment of the invention there is provided amethod for the treatment of advanced (metastatic) estrogen and/orprogesterone positive breast cancer in a warm-blooded animal, such asman, in need of such treatment, which comprises administering to saidanimal an effective amount of Compound (I) difumarate in combinationwith an effective amount of an aromatase inhibitor such as anastrozole,wherein said animal has not previously been treated with an endocrinetherapy such as for example, a selective estrogen receptor modulatorsuch as tamoxifen, an aromatase inhibitor such as anastrozole or anestrogen receptor down-regulator.

In another embodiment of the invention there is provided a method forthe treatment of non-metastatic estrogen and/or progesterone positivebreast cancer in a warm-blooded animal, such as man, in need of suchtreatment, which comprises administering to said animal an effectiveamount of Compound (I) difumarate in combination with an effectiveamount of an aromatase inhibitor such as anastrozole, wherein saidanimal has not previously been treated with an endocrine therapy such asfor example, a selective estrogen receptor modulator such as tamoxifen,an aromatase inhibitor such as anastrozole or an estrogen receptordown-regulator. In this embodiment the combination is suitablyadministered as an adjuvant treatment.

In the above two embodiments for the treatment of breast cancer, thewarm-blooded animal is suitably a post-menopausal woman. The term“post-menopausal” includes women that are naturally post-menopausal andwomen where the menopause has been induced, by for example, treatmentwith an LHRH agonist such as goserelin. It is to be understood thatwhere herein it is stated that a patient “not previously been treatedwith an endocrine therapy”, it is intended that the treatment of apatient with an LHRH agonist to induce early menopause in the patient isnot considered to be “previously treated with an endocrine therapy”.Accordingly, patients that have been treated with an LHRH agonist toinduce early menopause are not excluded from those embodiments that aredescribed herein as not being “treated with an endocrine therapy”.

In another embodiment the Compound (I) difumarate is used in combinationwith a taxane such as paclitaxel or docetaxel. This combination may beuseful in the treatment of breast cancer. For example, in the treatmentof a breast cancer (particularly advanced/metastatic breast cancer)which has a low over-expression of erbB2. The term “low over-expressionof erbB2” refers to tumours that are Her2 fluorescent in-situhybridization (FISH) negative. Particular tumours that are “lowover-expression of erbB2” those that are:

(i) Her2+ by immunohistochemistry (IHC); and/or

(ii) Her2++ by IHC and Her2 fluorescent in-situ hybridization (FISH)negative.

Accordingly in a particular embodiment of the invention the Compound (I)difumarate is used in combination with a taxane such as paclitaxel ordocetaxel in the is treatment of a cancer with low over-expression oferbB2 selected from one or more of:

(a) a breast cancer which is Her2 FISH negative;

(b) a breast cancer which is Her2+ by IHC; and

(c) a breast cancer which is Her2++ by IHC and Her2 FISH negative.

According to a further aspect of the present invention there is provideda method for the treatment of breast cancer with low over-expression oferbB2 in a warm-blooded animal, such as man, in need of such treatment,which comprises administering to said animal an effective amount ofCompound (I) difumarate in combination with an effective amount of ataxane such as paclitaxel or docetaxel.

Herein, where the term “combination” is used it is to be understood thatthis refers to simultaneous, separate or sequential administration. Inone aspect of the invention “combination” refers to simultaneousadministration. In another aspect of the invention “combination” refersto separate administration. In a further aspect of the invention“combination” refers to sequential administration. Where theadministration is sequential or separate, the delay in administering thesecond component should not be such as to lose the beneficial effect ofthe combination.

As will be understood the references to the use of Compound (I)difumarate described in the methods, uses and pharmaceuticalcompositions described herein, refer to any of the difumarates describedherein, for example Form A.

EXAMPLES

The invention is further illustrated by way of the following examples,which are intended to elaborate several embodiments of the invention.These examples are not intended to, nor are they to be construed to,limit the scope of the invention. It will be clear that the inventionmay be practiced otherwise than as particularly described herein.Numerous modifications and variations of the present invention arepossible in view of the teachings herein and, therefore, are within thescope of the invention.

In the Examples unless otherwise stated:

(i) yields are given for illustration only and are not necessarily themaximum attainable;

(ii) melting points were determined by DSC analysis using a MettlerDSC820e apparatus; 1-2 mg samples were accurately weighed and analysedin a vented sample pan; heating was carried out at 10° C./minute from25° C. to 325° C.; unless states otherwise melting points herein referto the onset temperature of the melting endotherm measured using DSC;

(iii) mass spectra were run with an electron energy of 70 electron voltsin the chemical ionization (CI) mode using a direct exposure probe;where indicated ionization was effected by electron impact (EI), fastatom bombardment (FAB) or electrospray (ESP); values for m/z are given;generally, only ions which indicate the parent mass are reported;

and unless otherwise stated, the mass ion quoted is (MH)⁻ which refersto the protonated mass ion; reference to M⁺ is to the mass ion generatedby loss of an electron; and reference to M−H⁺ is to the mass iongenerated by loss of a proton;

(iv) when given, NMR data is in the form of delta values for majordiagnostic protons, given in parts per million (ppm) relative totetramethylsilane (TMS) as an internal standard, determined at 500 MHzusing perdeuterio dimethyl sulfoxide (DMSO-d₆) as solvent unlessotherwise indicated; the following abbreviations have been used: s,singlet; d, doublet; t, triplet; q, quartet; m, multiplet; br, broad;

(v) chemical symbols have their usual meanings; SI units and symbols areused;

(vi) solvent ratios are given in volume:volume (v/v) terms;

(vii) thermogravimetric analysis was carried out using Mettler TG851equipment [1-5 mg samples were accurately weighed and analysed in anopen pan; heating was carried out at 10° C./minute from 25° C. to 325°C.

(viii) X-Ray Powder Diffraction analysis was carried out using a SiemensD5000 powder X-ray diffractometer fitted with a scintillation detector;the X-Ray source was Cu K_(α), giving a wavelength of 1.54 Å; data werecollected over the range 2-theta 2-40°, in increments of 2-theta 0.02°,with 1 second per increment and was categorised into the categoriesidentified in Table 2 below:

TABLE 2 % Relative Intensity* Definition  25-100 vs (very strong) 10-25s (strong)  3-10 m (medium) 1-3 w (weak) *The relative intensities arederived from diffractograms measured with fixed slits

[As previously stated, persons skilled in the art of X-ray powderdiffraction will realise that the relative intensity of peaks can beaffected by, for example, grains above 30 microns in size andnon-unitary aspect ratios, which may affect analysis of samples. Theskilled person will also realise that the position of reflections can beaffected by the precise height at which the sample sits in thediffractometer and the zero calibration of the diffractometer. Thesurface planarity of the sample may also have a small effect. Hence thediffraction pattern data presented are not to be taken as absolutevalues (see Jenkins, R & Snyder, R. L. ‘Introduction to X-Ray PowderDiffractometry’ John Wiley & Sons, 1996, for further information)];

(ix) dynamic vapour sorption was measured using a SMS DVS (SurfaceMeasurement Systems Limited, UK). Samples were analysed at 25° C. usinga gas flow of 200 cubic centimetres per minute. The relative humidity(RH) was increased from 0% RH in steps of 10% RH to 80% RH with thefinal step of 95% RH. The sample was then desorbed using the same RHstep pattern as the sorption; this procedure was then repeated in asecond sorption/desorption cycle. Equilibration at each humidity step isset such that the rate of change of weight with time (minute) was0.002%.

(x) intrinsic dissolution rate was measured in a dissolution bathcoupled to a fibre optic uv detector.

(xi) solubility in water was measured using HPLC UV.

(xii) in the examples given below the number of moles and the yieldstated refer to the raw materials and reagents at 100% w/w, therebytaking account of the purity of the materials used.

Example A Preparation of4-(3-Chloro-2-fluoroanilino)-7-methoxy-6-{[1-(N-methylcarbamoylmethyl)piperidin-4-yl]oxy}quinazoline(Compound (I))

2-Chloro-N-methylacetamide (3.720 kg, 34.60 mol) and4-(3-chloro-2-fluoroanilino)-7-methoxy-6-[(piperidin-4-yl)oxy]quinazolinedihydrochloride (13.70 kg, 27.25 mol), were dissolved in acetonitrile(79.2 kg). To the stirred suspension, at ambient temperature, was addedtriethylamine (17.40 kg, 172.11 mol). The resulting clear solution washeated to reflux and held for 3 hours. The solution was cooled to 20° C.(product crystallized at 50° C.). Water (54.2 kg) was added to thereactor and the suspension was stirred for a further 2 hours at 20° C.The product was filtered and washed with water (34 kg) followed by cold(0° C.) acetonitrile (13.0 kg). The product was recrystallised fromacetonitrile (94.6 kg), isolated by filtration and washed with cold (0°C.) acetonitrile (13.2 kg). A further recrystallisation of this productwas then carried out as above from acetonitrile (75.2 kg). The solid wasthen dried under vacuum to give the title product as a white solid (6.50kg, 50%); ¹H NMR Spectrum: (CDCl₃) 1.98 (m, 2H), 2.08 (m, 2H), 2.46 (-m,2H), 2.85 (m, 2H), 2.87 (d, 3H), 3.07 (s, 2H), 4.02 (s, 3H), 4.49 (m,1H), 7.16 (m, 4H), 7.31 (m, 2H), 8.49 (m, 1H), 8.71 (s, 1H); Massspectrum: MH⁻ 474.

The4-(3-Chloro-2-fluoroanilino)-7-methoxy-6-[(piperidin-4-yl)oxy]quinazolineused as the starting material was prepared as follows:

Step 1: 6-Acetoxy-4-(3-chloro-2-fluoroanilino)-7-methoxyquinazolinehydrochloride

6-Acetoxy-7-methoxyquinazolin-4-one (International Patent Application WO96/15118, Example 39 thereof; 21.4 kg, 89.3 mol) was suspended intoluene (150 kg). To this was added N-ethyldiisopropylamine (13.3 kg,103 mol). The brown suspension was heated to 70° C. then phosphorusoxychloride (36.0 kg, 228 mol) was charged. The reaction mixture wasstirred at 70° C. for 5 hours. Further toluene (84.0 kg) was addedfollowed by 3-chloro-2-fluoroaniline (14.88 kg, 102 mol). The reactionmixture was stirred at 70° C. for 2 hours during which time a solidprecipitated. The suspension was cooled to 25° C. and held at thistemperature for 93 hours. The reaction mixture was filtered and thefilter cake washed with toluene (2×55.5 kg). The cake was further washedwith a mixture of ethanol (24.5 kg) and water (32.0 kg) twice, thenethanol (50.5 kg) twice and the solid then dried under vacuum to givethe title product as a beige solid (33.4 kg, 78%); ¹H NMR: 2.37 (s, 3H),4.00 (s, 3H), 7.34 (ddd, 1H), 7.48 (s, 1H), 7.52 (ddd, 1H), 7.61 (ddd,1H), 8.62 (s, 1H), 8.86 (s, 1H); Mass Spectrum: 362.4, 364.4.

Step 2: 4-(3-Chloro-2-fluoroanilino)-6-hydroxy-7-methoxyquinazoline

6-Acetoxy-4-(3-chloro-2-fluoroanilino)-7-methoxyquinazolinehydrochloride from step 1 (33.5 kg, 69.6 mol) was suspended in methanol(198 kg). To the stirred suspension at 25° C. was added water (86 kg)and sodium hydroxide (31.5 kg, 32%). The resulting solution was stirredat 60° C. for 4.5 hours and then cooled to 25° C. Acetic acid(approximately 16.0 kg) was added until a pH of 5.5-6.0 was achieved atwhich point the product precipitates from solution. After the additionof further methanol (5.5 kg) the suspension was stirred for 90 minutes.The product was filtered then washed with 25% aqueous methanol (39.0 kgMeOH+17.0 kg Water) and then methanol (55.5 kg). The crude solid wasdried under vacuum at 40° C. The crude solid was slurried with water(145 kg) and stirred for 2 hours at 65° C. The slurry was cooled to 20°C. and filtered. The filter cake was washed with methanol (2×21.5 kg),then dried under vacuum at 40° C. to give a the title product as a lightbrown solid (21.85 kg, 98%); ¹H NMR: 3.95 (s, 3H), 7.19 (s, 1H), 7.23(dd, 1H), 7.42 (dd, 1H), 7.50 (dd, 1H), 7.64 (s, 1H), 8.32 (s, 1H),9.43(s, 1H), 9.67 (br.s, 1H); Mass Spectrum: 320.4, 322.4.

Step 3:6-{[1-tert-Butoxycarbonyl)piperidin-4-yl]oxy}-4-(3-chloro-2-fluoroanilino)-7-methoxyquinazoline

4-(3-Chloro-2-fluoroanilino)-6-hydroxy-7-methoxyquinazoline from Step 2(15.591 kg, 48.44 mol), tent-Butyl(4-methanesulfonyloxy)piperidine-1-carboxylate (prepared as in Chemical& Pharmaceutical Bulletin 2001, 49(7), 822-829; 16.20 kg, 57.99 mol) andpotassium carbonate (7.978 kg, 57.73 mol) were dissolved inN-methylpyrrolidinone (114.2 kg), and the mixture was heated to 100° C.with stirring. Heating was continued at 100° C. (95° C.-105° C.) for 5hours. The mixture was then cooled to 80° C. and quenched by theaddition of water (216.6 kg).

The batch was stirred at 80° C. for a further 60 minutes then cooled to20° C. over 2 hours, during which time the product crystallized. Theproduct was isolated by filtration. The product was dissolved in hot(reflux) methanol (200 L). To this mixture was added water (20 L), whichinduced crystallization. The suspension was cooled to 0° C. andfiltered. Vacuum drying at 50° C. afforded the title product, 18.80 kg(77%); ¹H NMR: 1.40 (s, 9H), 1.60-1.65 (m, 2H), 1.95-2.00 (m, 2H),3.20-3.25 (m, 2H), 3.65-3.70 (m, 2H), 3.92 (s, 3H), 4.68 (m, 1H), 7.21(s, 1H), 7.27 (dd, 1H), 7.47 (ddd, 1H), 7.51 (dd, 1H), 7.85 (s, 1H),8.36 (s, 1H), 9.53 (s, 1H); Mass Spectrum: 503.5, 505.5.

Step 4:4-(3-chloro-2-fluoroanilino)-7-methoxy-6-[(piperidin-4-yl)oxy]quinazolinedihydrochloride

6-{[(1-tert-Butoxycarbonyl)piperidin-4-yl]oxy}-4-(3-chloro-2-fluoroanilino)-7-methoxyquinazoline from step 3 (18.80 kg,37.38 mol) was suspended in isopropanol (139.8 kg), and heated to 40° C.with stirring. Hydrochloric acid (15.40 kg, 156.3 mol) was charged tothe vessel over 50 minutes, allowing an exotherm of approximately 9° C.to occur. During the charging of the acid, the suspension dissolved togive a clear solution. The solution was heated slowly to reflux overapproximately 90 minutes, and then held at reflux for a further 3 hours.The product crystallised out during this reflux period. The thicksuspension was cooled to 0° C. and filtered. The filter cake was washedtwice with cold (0° C.) isopropanol (2×20.6 kg). The product was driedunder vacuum at 50° C. to give the title product, 13.60 kg (73%); ¹HNMR: 1.53-1.64 (m, 2H), 2.00-2.05 (m, 2H), 2.64-2.72 (m, 2H), 3.00-3.07(m, 2H), 3.92 (s, 3H), 4.60 (m, 1H), 7.20 (s, 1H), 7.26 (dd, 1H), 7.47(dd, 1H), 7.50 (dd, 1H), 7.82 (s, 1H), 8.34 (s, 1H), 9.56 (s, 1H); MassSpectrum: 403.2, 405.2.

Example B Preparation of4-(3-Chloro-2-fluoroanilino)-7-methoxy-6-{[1-(N-methylcarbamoylmethyl)piperidin-4-yl]oxy}quinazoline(Compound (I))

2-Chloro-N-methylacetamide (24.22 g, 223.1 mmol) and4-(3-chloro-2-fluoroanilino)-7-methoxy-6-[(piperidin-4-yl)oxy]quinazolinedihydrochloride (86.00 g, 160.9 mmol), were slurried in acetonitrile(537 ml). To the stirred suspension, at ambient temperature, was addedtriethylamine (101 ml, 723.9 mmol). The reaction was heated to 75° C.held for 5 hours. The solution was cooled to 70° C. and ethanol (268 ml)added. The reaction was cooled to 45° C. and water (9.6 ml) added.Compound (I) (0.42 g) was added to establish crystallisation and thenthe slurry cooled to 20° C. over 2 hours. After stirring for a further12 hours the product was isolated by filtration. The filter cake waswashed twice with acetonitrile (102 ml): ethanol (51 ml): water (1.8 ml)and then with water (153 ml). The product was dried in vacuo at 60° C.to give the title compound as a white solid (45.9g, 60%); 1H NMR (400MHz, DMSO-d₆) δ ppm 1.76-1.87 (m, 2H) 2.01 -2.11 (m, 2H) 2.35 -2.44 (m,2H) 2.64 (d, J=4.74 Hz, 3H) 2.72 -2.80 (m, 2H) 2.95 (s, 2H) 3.95 (s, 3H)4.51-4.63 (m, 1H) 7.23 (s, 1H) 7.29 (td, J=8.08, 1.29 Hz, 1H) 7.46-7.58(m, 2H) 7.75 (q, J=4.60 Hz, 1H) 7.83 (s, 1H) 8.38 (s, 1H) 9.59 (s,1H)Mass spectrum: MH⁺474.

The4-(3-chloro-2-fluoroanilino)-7-methoxy-6-[(piperidin-4-yl)oxy]quinazolinedihydrochloride used as the starting material was prepared as follows:

Step 1:6-{[(1-tert-Butoxycarbonyl)piperidin-4-yl]oxy}-4-(3-chloro-2-fluoroanilino)-7-methoxyquinazoline

4-(3-Chloro-2-fluoroanilino)-6-hydroxy-7-methoxyquinazoline (prepared asdescribed in Step 2 of Example A; 60.00 g, 0.1828 mol), tent-Butyl(4-methanesulfonyloxy)piperidine-1-carboxylate (88.04 g, 0.3107 mol) andpotassium carbonate (30.31 g, 0.2193 mol) were suspended in ethanol (584ml) and water (58 ml), and the mixture was heated to reflux withstirring. Heating was continued at reflux for 16.5 hours. The mixturewas then cooled to 70° C. and water (234 ml) was added over 60 minutes.

The batch was stirred at 65° C. for a further 2 hours to establishcrystallisation. The slurry was cooled to 20° C. over 6 hours. Theproduct was isolated by filtration. The filter cake was slurried withaqueous ethanol (ethanol 117 ml, water 58 ml) and then displacementwashed with aqueous ethanol (ethanol 117 ml, water 58 ml). The filtercake was then slurried with water (175 ml) and then displacement washedwith water (175 ml). The product was dried in vacuo at 40° C. to givethe title compound (81.5 g, 84%); 1H NMR (500 MHz, DMSO-d₆) δ ppm 1.42(s, 9H) 1.60-1.70 (m, 2H) 1.96-2.04 (m, 2H) 3.23-3.30 (m, 2H) 3.65-3.75(m, 2H) 3.95 (s, 3H) 4.68-4.75 (m, 1H) 7.24 (s, 1H) 7.29 (t, J=8.06 Hz,1H) 7.49 (t, J=7.50 Hz, 1H) 7.54 (t, J=7.19 Hz, 1H) 7.88 (s, 1H) 8.39(s, 1H) 9.57 (s, 1H); Mass Spectrum: 503.5, 505.5.

Step 2:4-(3-chloro-2-fluoroanilino)-7-methoxy-6-[(piperidin-4-yl)oxy]quinazolinedihydrochloride

6-{[(1-tert-Butoxycarbonyl)piperidin-4-yl]oxy}-4-(3-chloro-2-fluoroanilino)-7-methoxyquinazoline(10.00 g, 0.1879 mol) was suspended in industrial methylated spirits (95ml), and heated to 35° C. with stirring. Hydrochloric acid (6.59 ml,approximately 0.7891 mol) was charged to the vessel allowing an exothermof approximately 5.5° C. to occur. During the charging of the acid, thesuspension dissolved to give a clear solution. The solution was heatedslowly to 70° C. over approximately 90 minutes, and then held at 70° C.for a further 1 hour. The reaction is then cooled to 0° C. over 4 hoursduring which time the product crystallises. The product was isolated byfiltration and then filter cake was washed twice with industrialmethylated spirits (2×14 ml). The product was dried in vacuo at 50° C.to give the title product (9.04 g, 88%); 1H NMR (400 MHz, DMSO-d₆) δ ppm1.91-2.01 (m, 2H) 2.27-2.35 (m, 2H) 3.15-3.26 (m, 2H) 3.26-3.35 (m, 2H)4.02 (s, 3H) 5.07-5.15 (m, 1H) 7.35 (td, J=8.08, 1.29 Hz, 1H) 7.46 (s,1H) 7.52 (ddd, J=8.03, 5.23 Hz, 1H) 7.63 (ddd, J=8.22, 6.76, 1.62 Hz,1H) 8.83 (s, 1H) 8.91 (s, 1H) 9.02-9.13 (m, 1H) 9.20-9.31 (m, 1H) 12.51(br. s., 1H)); Mass Spectrum: 403.2, 405.2.

Example C Preparation of4-(3-Chloro-2-fluoroanilino)-7-methoxy-6-{[1-(N-methylcarbamoylmethyl)piperidin-4-yl]oxy}quinazoline(Compound (I))

Compound (I) was prepared according to the scheme shown below:

2-[4-(5-cyano-4-{[(dimethylamino)methylene]amino}-2-methoxyphenoxy)piperidin-1-yl]-N-methylacetamide(7, 7.00 g, 17.71 mmoles), was suspended in methoxybenzene (35.8 g).Acetic acid (16.6 g) was charged and to the resulting solution was added3-chloro-2-fluoroaniline (2.71 g, 18.07 mmoles). The reaction mixturewas heated at 90° C. for 20 hours then cooled to 20° C. Water (37.04 g)was charged to the reaction mixture, and the organic layer discarded. Tothe resulting aqueous mixture was charged isopropanol (39.00 g),followed by aqueous ammonia (20.79 g, 25%). The reaction mixture washeated to 30° C. and seeded with Compound (I), which inducedcrystallisation. The reaction was then cooled to 0° C. and the productisolated by filtration. The filter cake was washed twice with a mixtureof water (7.28 g) and isopropanol (4.68 g), then dried to afford theCompound (I) (5.65 g, 55% yield); ¹H NMR (400 MHz, DMSO-d₆) δ ppm 1.79(m, 2H) 2.04 (m, 2H) 2.38 (m, 2H) 2.62 (d, J=4.5 Hz, 3H) 2.74 (m, 2H)2.94 (s, 2H) 3.93 (s, 3H) 4.56 (tt, J=8.1, 3.8 Hz, 1H) 7.21 (s, 1H) 7.28(m, 1H) 7.50 (m, 2H) 7.73 (q, J=4.5 Hz, 1H) 7.81 (s, 1H) 8.36 (s, 1H)9.56 (br.s, 1H); Mass Spectrum: m/z (M +H)⁺ 474.2, 476.2.

The2-[4-(5-cyano-4-{[(dimethylamino)methylene]amino}-2-methoxyphenoxy)piperidin-1-yl]-N-methylacetamide(7), used as the starting material was prepared as follows:

Step 1. Preparation of tert-butyl4-(5-cyano-2-methoxyphenoxy)piperidine-1-carboxylate (2)

3-hydroxy-4-methoxybenzonitrile (1, 6.00 g, 39.62 mmole), tert-butyl(4-methanesulfonyloxy)piperidine-1-carboxylate (16.6 g, 59.44 mmoles)(Chemical & Pharmaceutical Bulletin 2001, 49(7), 822-829); and potassiumcarbonate (6.71 g, 47.55 mmoles) were suspended in isopropanol (78.98 g)and the mixture was heated at reflux with stirring. Additionaltert-butyl (4-methanesulfonyloxy)piperidine-1-carboxylate (2.08 g, 7.43mmoles) was added to push the reaction to completion. The mixture wasthen cooled and quenched by the addition of water (100.47 g). Seedingwith tert-butyl 4-(5-cyano-2-methoxyphenoxy)piperidine-1-carboxylate (2)followed by cooling to 0° C. resulted in a crystalline product, whichwas isolated by filtration. The filter cake was washed with a mixture ofwater (8.86 g) and isopropanol (6.97 g), followed by water (23.64 g) andthen dried to give the title compound (10.75 g, 80% yield); ¹H NMR (400MHz, DMSO-d₆) δ ppm 1.39 (s, 9H) 1.48 (m, 2H) 1.88 (m, 2H) 3.13 (m, 2H)3.67 (m, 2H) 3.83 (s, 3H) 4.56 (tt, J=8.1, 3.8 Hz, 1H) 7.13 (d, J=8.4Hz, 1H) 7.42 (dd, J=8.4, 1.9 Hz, 1H) 7.51 (d, J=1.9 Hz, 1H); MassSpectrum: m/z (M+H)⁺ 333.1.

Step 2. Preparation of 4-methoxy-3-(piperidin-4-yloxy)benzonitrile (3)

Tert-butyl 4-(5-cyano-2-methoxyphenoxy)piperidine-1-carboxylate (2,39.31 g, 118.26 mmoles) was suspended in ethanol (155.53 g) and heatedto 40° C. To this slurry was slowly added HCl (46.61 g, 573.04 mmoles).The mixture was heated to 60° C. and held for 3 hours. The reactionmixture was cooled to 20° C. and seed was charged initiatingcrystallisation. The resulting solid was isolated by filtration at 0°C., washed twice with ethanol (62.21 g) and then dried to give the titlecompound as the hydrochloride salt (29.84 g, 77% yield); ¹H NMR (400MHz, DMSO-d₆) δ ppm 1.84 (m, 2H) 2.09 (m, 2H) 3.02 (ddd, J=12.7, 8.9,3.4 Hz, 2H) 3.20 (m, 2H) 3.84 (s, 3H) 4.63 (tt, J=7.7, 3.6 Hz, 1H) 7.15(d, J=8.5 Hz, 1H) 7.45 (dd, J=8.5, 1.9 Hz, 1H) 7.56 (d, J=1.9 Hz, 1H)9.16 (br. s, 2H); Mass Spectrum: m/z (M+H)⁺ 233.2.

Step 3. Preparation of2-[4-(5-cyano-2-methoxyphenoxy)piperidin-1-yl]-N-methylacetamide (4)

4-Methoxy-3-(piperidin-4-yloxy)benzonitrile hydrochloride salt (3, 28.36g, 95.82 mmoles), 2-chloro-N-methylacetamide (12.37 g, 114.98 mmoles)and potassium carbonate (33.11 g, 239.55 mmoles) were suspended inacetonitrile (161.36 g). The reaction mixture was heated at reflux for 3hours. The reaction mixture was cooled to 20° C. and water (386.26 g)was charged. The reaction was heated to 75° C. and the volume reduced bydistillation. Upon cooling crystallisation occurred. The resulting solidwas isolated by filtration, washed twice with water (77.25 g and 128.75g) and then dried to give the title is compound (27.95 g, 94% yield); ¹HNMR (400 MHz, DMSO-d₆) δ ppm 1.68 (m, 2H) 1.91 (m, 2H) 2.29 (m, 2H) 2.61(d, J=4.7 Hz, 3H) 2.67 (m, 2H) 2.88 (s, 2H) 3.83 (s, 3H) 4.41 (tt,J=8.3, 4.0 Hz, 1H) 7.11 (d, J=8.4 Hz, 1H) 7.40 (dd, J=8.4, 1.9 Hz, 1h)7.47 (d, J=1.9 Hz, 1H) 7.68 (q, J=4.7 Hz, 1H); Mass Spectrum: m/z (M+H)⁺ 304.2.

Step 4. Preparation of2-[4-(5-cyano-2-methoxy-4-nitrophenoxy)piperidin-1-yl]-N-methylacetamide(5)

2-[4-(5-Cyano-2-methoxyphenoxy)piperidin-1-yl]-N-methylacetamide (4,8.78 g, 26.11 mmoles) was suspended in acetic acid (22.82 g, 364.87mmoles) and the resulting reaction mixture cooled to 5° C. To this wasadded sulfuric acid (23.64 g, 234.95 mmoles) maintaining the reactiontemperature below 30° C. To the resulting solution was added nitric acid(2.40 g, 26.63 mmoles). The reaction mixture was then heated to 35° C.and held for 3 hours. Additional nitric acid (117 mg, 1.31 mmoles) andsulphuric acid (1.31 g 13.1 mmoles) were charged and the reactionmixture was heated at 35° C. for 30 minutes. The solution was cooled to20° C. and quenched with aqueous ammonia (92.45 g 1.36 moles), resultingin an increase in temperature to 50° C. To the resulting slurry wasadded, propionitrile (61.58 g 1.12 moles) and water (19 g). The reactionmixture was heated to 80° C. resulting in a clear solution, which uponsettling gave two layers. The bottom layer was removed. The reactionmixture was cooled to 20° C. resulting in a thick slurry. The solid wasisolated by filtration, washed with propionitrile (6.16 g 112.0 mmoles)and dried to afford the title compound (7.44 g, 82% yield); ¹H NMR (400MHz, DMSO-d₆) δ ppm 1.72 (m, 2H) 1.97 (m, 2H) 2.35 (m, 2H) 2.61 (d,J=4.7 Hz, 3H) 2.66 (m, 2H) 2.90 (s, 2H) 3.96 (s, 3H) 4.73 (tt, J=8.4,4.0 Hz, 1H) 7.71 (q, J=4.7 Hz, 1H) 7.82 (s, 1H) 7.86 (s, 1H). MassSpectrum: m/z (M+H)⁺ 349.2

Step 5. Preparation of2-[4-(4-amino-5-cyano-2-methoxyphenoxy)piperidin-1-yl]-N-methylacetamide(6)

2-[4-(5-Cyano-2-methoxy-4-nitrophenoxy)piperidin-1-yl]-N-methylacetamide(5, 7.42 g, 19.38 mmoles) was suspended in water (44.52 g) and methanol(5.35 g). To this was added sodium dithionite (11.91 g, 58.15 mmoles)and the resulting reaction mixture was heated to 60° C. To the reactionmixture was added hydrochloric acid (46.98 g, 463.89 mmoles)), resultingin a solution, which was held at 60° C. for 3 hours. The reactionmixture was then allowed to cool to 20° C. Aqueous sodium hydroxide(15.51 g 182.2 mmoles) was charged followed by 2-methyltetrahydrofuran(58.0 g). The reaction mixture was heated to 60° C., which upon settlinggave two layers and the lower aqueous layer was is discarded. The volumeof the reaction mixture was reduced by vacuum distillation and methyltert-butyl ether (18.54 g) was added to give a slurry which was cooledto 10° C. and then the solid was collected by filtration. The solid waswashed with 2-methyltetrahydrofuran (5.8 g) and dried to give the titlecompound (5.4 g, 78% yield); ¹H NMR (400 MHz, DMSO-d₆) δ ppm 1.62 (m,2H) 1.82 (m, 2H) 2.20 (m, 2H) 2.60 (d, J=4.7 Hz, 3H) 2.65 (m, 2H) 2.86(s, 2H) 3.72 (s, 3H) 4.00 (tt, J=8.3, 4.0 Hz, 1H) 5.66 (br. s, 2H) 6.39(s, 1H) 6.94 (s, 1H) 7.65 (q, J=4.7 Hz, 1H)

Mass Spectrum: m/z (M +H)⁺ 319.2

Step 6. Preparation of2-[4-(5-cyano-4-{[(dimethylamino)methylene]amino}-2-methoxyphenoxy)piperidin-1-yl]-N-methylacetamide(7)

2-[4-(4-Amino-5-cyano-2-methoxyphenoxy)piperidin-1-yl]-N-methylacetamide(6, 18.21 g, 52.05 mmoles) was suspended in 2-methyltetrahydrofuran(99.62 g). To this was added acetic acid (162.79 mg), andN,N-dimethylformamide dimethyl acetal (DMF-DMA) (8.63 g, 70.27 mmoles)and the resulting reaction mixture was heated at 76° C. for 16 hrs.Additional N,N-dimethylformamide dimethyl acetal (639.41 mg, 5.20mmoles) was added to the reaction mixture to ensure the reactioncompleted. The reaction mixture was cooled to 30° C. during which timecrystallisation occurred. The resulting solid was isolated byfiltration, washed with 2-methyltetrahydrofuran (14.23 g) and dried toafford the title compound (19.53 g, 97% yield); ¹H NMR (400 MHz,DMSO-d₆) δ ppm 1.65 (m, 2H) 1.86 (m, 2H) 2.24 (m, 2H) 2.60 (d, J=4.7 Hz,3H) 2.66 (m, 2H) 2.87 (s, 2H) 2.95 (s, 3H) 3.04 (s, 3H) 3.81 (s, 3H)4.19 (tt, J=8.2, 3.8 Hz, 1H) 6.72 (s, 1H) 7.15 (s, 1H) 7.67 (q, J=4.7Hz, 1H) 7.90 (s, 1H); Mass Spectrum: m/z (M+H)⁺ 374.2.

Biological Activity

The activity of Compound (I) was to assessed to test its ability to:

a) inhibit the activation (phosphorylation) of EGFR, ErbB2 and ErbB3 inligand-stimulated cells; and

b) inhibit the basal and ligand-stimulated proliferation of MCF-7 cells.

(a) Compound (I) in Ligand Driven Assays Methods:

KB cells and MCF-7 cells were obtained from the American Type CultureCollection (ATCC) and routinely cultured in RPMI 1640 (Phenol redfree)+10% Foetal Bovine Serum+2 mM L-Glutamine.

Treatment and Lysis of Cells:

KB cells were seeded at 5000 cells/well and MCF-7 cells at4000cells/well in 96 well plates in RPMI 1640 media containing 10% FBS.Cells were incubated for 72 hours before replacing the media withserum-free RPMI 1640 media for 24 hours. Cells were then treated withCompound (I) for 90 minutes at concentrations ranging from 0-10 μM.Immediately before cell lysis, MCF-7 and KB cells were incubated for 5minutes with ligand (heregulin (“HRG”) for the MCF-7 cells and epidermalgrowth factor (“EGF”) for the KB cells) at concentrations required toincrease receptor phosphorylation to 90% of max (ED₉₀) to allowinter-assay comparison.

Measurement of p-EGFR, p-ErbB2 and p-ErbB3:

The p-EGFR status of KB cells was measured using the Human phospho-EGFRDuoset ELISA kit (R&D systems total EGFR #DYC1854, pEGFR #DYC1095). Thep-ErbB2 and p-ErbB3 content of MCF-7 cells were measured using the Humanphospho-ErbB2 Duoset ELISA kit (R&D systems, DYC1768) and Humanphospho-ErbB3 Duoset ELISA kit (R&D systems, DYC1769) respectively. Thekits measured whole cell tyrosine phosphorylation of EGFR, ErbB2 orErbB3. Assays were performed according to the manufacturersinstructions, with 50 μl lysate added per well.

Results:

The results are summarised in Table 3

TABLE 3 Compound (I) activity against p-EGFR (in KB cells) and p-ErbB2and p- ErbB3 (in MCF-7 cells) p-EGFR p-ErbB2 p-ErbB3 Geo Mean IC₅₀ GeoMean IC₅₀ Geo Mean IC₅₀ Compound (95% CIR*) (95% CIR) (95% CIR) Compound(I) 0.004 (1.377) 0.003 (1.817) 0.004 (1.89) *Confidence Interval Ratio

Table 3 shows that Compound (I) is a potent inhibitor of phospho-EGFR,phospho-ErbB2 and phospho-ErbB3 in these cells.

b) Compound (I) in Basal or HRG-Stimulated MCF-7 Cell ProliferationAssay Methods:

MCF-7 cells were routinely cultured in DMEM (Phenol red free)+10% FoetalBovine Serum+2 mM L-Glutamine.

Cells were seeded at 4000 cells per well in 96 well plates in DMEM mediacontaining 1% charcoal/dextran-treated FBS and 2 mM glutamine andallowed to settle for 4 hours prior to treatment with Compound (I) atconcentrations ranging from 0-3 μM and is 0-10 μg/ml respectively. Twohours following treatment, cells were incubated with 10 ng/ml HRG, aconcentration required to increase MCF-7 cell proliferation to 90% ofmax (ED₉₀). Basal wells were unstimulated with ligand. After incubationfor 4 days, cell viability was assessed using a3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium(MTS) assay.

Prior to IC₅₀ determination of HRG-stimulated compound-treated cells,mean basal growth at 96 hours was subtracted from each of the readoutsso that proliferation driven through HRG-signalling was assessed. BasalIC₅₀ values are expressed as GI₅₀ i.e. the day 0 plate cell number(baseline reading) was subtracted from the readout at 96 hours later.

Results:

The results are summarised in Tables 4 and 5.

TABLE 4 HRG-stimulated proliferation IC₅₀ value Compound Geo Mean IC₅₀95% CIR Compound (I) 0.061 μM 2.421

TABLE 5 Basal proliferation GI₅₀ values Compound Geo Mean GI₅₀ 95% CIRCompound (I) 1.094 μM 4.423

In the KB cells, stimulation with EGF, which specifically binds to EGFR,causes phosphorylation and therefore activation of this receptor.Similarly in the MCF-7 cells HRG, which binds specifically to ErbB3causes it to form heterodimers with ErbB2 and both receptors becomephosphorylated and activated. Tables 4 shows that Compound (I) is apotent inhibitor of HRG-stimulated MCF-7 proliferation. These effects onproliferation are believed to be due to the activities of thesecompounds on ErbB2/ErbB3 heterodimers, with Compound (I) being a muchmore potent inhibitor of this heterodimer.

The MCF-7 basal assays represent a situation where there no increasedstimulation or activation of erbB2/erbB3 heterodimerisation. Table 5shows that even in such conditions Compound (I) inhibits MCF-7.

Example 1 Preparation of Compound (I) Difumarate Form A:2-[4-({4-1(3-Chloro-2-fluorophenyl)amino]-7-methoxyquinazolin-6-yl}oxy)piperidin-1-yl]-N-methylacetamidedi-[(2E)-but-2-enedioate] Form A

A solution of fumaric acid (2.7 g, 23.22 mmol) in methanol (95 ml) wasadded to a mixture of2-[4-({4-[(3-Chloro-2-fluorophenyl)amino]-7-methoxyquinazolin-6-yl}oxy)piperidin-1-yl]-N-methylacetamide(Compound (I)) (5.62 g at 89% w/w, 10.55 mmol) in isopropanol (100 ml)maintaining the temperature >65° C. The mixture was heated at reflux forone hour before clarification. The reaction mixture was cooled to 30° C.over 90 minutes and held for 30 minutes to establish crystallisation.The reaction was cooled to 0° C. over 2 hours and held for 1 hour beforeisolation by filtration. The filter cake was washed twice with coldisopropanol (2×10 ml) and dried in vacuo at 50° C. to give the titlecompound as a white solid (5.84 g, 78%); ¹H NMR Spectrum: (DMSO) 1.85(m, 1H), 2.08 (m, 1H), 2.50 (m, 1H), 2.66 (d, 3H), 2.83 (m, 1H), 3.05(s, 2H), 3.96 (s, 3H), 4.58 (m, 1H), 6.64 (s, 4H), 7.23 (s, 1H), 7.28(m, 1H), 7.46 (ddd, 1H), 7.55 (m, 1H), 7.70 (broad q, 1H), 7.85 (s, 1H),8.38 (s, 1H).

Example 2 Preparation of Compound (I) Difumarate Form A:2-[4-({4-[(3-Chloro-2-fluorophenyl)amino]-7-methoxyquinazolin-6-yl}oxy)piperidin-1-yl]-N-methylacetamidedi-[(2E)-but-2-enedioate] Form A

A solution of fumaric acid (1.4 kg, 12.1 mol) in methanol (26.6 kg) wasadded to a mixture of2-[4-({4-[(3-chloro-2-fluorophenyl)amino]-7-methoxyquinazolin-6-yl}oxy)piperidin-1-yl]-N-methylacetamide(2.93 kg, 84.8% w/w, 5.24 mol) in isopropanol (39 kg) maintaining thetemperature >65° C. A line wash of methanol (3.6 kg) was charged. Themixture was heated at reflux for one hour before clarification, followedby a line wash of methanol (7 kg). The reaction mixture was distilled atatmospheric pressure to remove 47 kg of distillates. Isopropanol (15.8kg was added and the reaction mixture distilled to remove 15.6 kg ofdistillates. Crystallisation occurred during the distillation.Isopropanol (21 kg) was added and the reaction cooled to 0° C. over 8hours and held for 1 hour before isolation by filtration. The filtercake was washed with cold 50:50 isopropanol:MeOH (4 kg) followed by coldisopropanol (4 kg) and dried in vacuo at 50° C. to give the titlecompound as a white solid (3.64 kg, 98%); ¹H NMR Spectrum: (DMSO) 1.85(m, 1H), 2.08 (m, 1H), 2.50 (m, 1H), 2.66 (d, 3H), 2.83 (m, 1H), 3.05(s, 2H), 3.96 (s, 3H), 4.58 (m, 1H), 6.64 (s, 4H), 7.23 (s, 1H), 7.28(m, 1H), 7.46 (ddd, 1H), 7.55 (m, 1H), 7.70 (broad q, 1H), 7.85 (s, 1H),8.38 (s, 1H).

Example 3 Preparation of Compound (I) Difumarate Form A:2-[4-({4-[(3-Chloro-2-fluorophenyl)amino]-7-methoxyquinazolin-6-yl}oxy)piperidin-1-yl]-N-methylacetamidedi-[(2E)-but-2-enedioate] Form A

2-[4-({4-[(3-Chloro-2-fluorophenyl)amino]-7-methoxyquinazolin-6-yl}oxy)piperidin-1-yl]-N-methylacetamide(Compound (I)) (60.19 g at 88% w/w, 111.8 mmol) was dissolved in ethylacetate (1550 ml). The solution was clarified by filtration and thefilter washed with ethyl acetate (53 ml). The solution was cooled to 40°C. A clarified solution of fumaric acid (26.60 g, 257.0 mmol) inisopropanol (408 ml) was then added over 1 hour. The filter used toclarify the fumaric acid solution was then washed with isopropanol (37ml). After holding for 1 hour at 40° C. the reaction was cooled to 20°C. over 1 hour. The reaction mixture was held for 13.5 hours beforeisolating the product by filtration. The filter cake was washed twicewith ethyl acetate (82 ml): isopropanol (24 ml) and then dried in vacuoat 40° C. to give the title compound as a white solid (72.32 g, 90%); ¹HNMR Spectrum: (DMSO) 1.85 (m, 1H), 2.08 (m, 1H), 2.50 (m, 1H), 2.66 (d,3H), 2.83 (m, 1H), 3.05 (s, 2H), 3.96 (s, 3H), 4.58 (m, 1H), 6.64 (s,4H), 7.23 (s, 1H), 7.28 (m, 1H), 7.46 (ddd, 1H), 7.55 (m, 1H), 7.70(broad q, 1H), 7.85 (s, 1H), 8.38 (s, 1H).

Example 4 Preparation of Compound (I) Difumarate Form A:2-[4-({4-[(3-Chloro-2-fluorophenyl)amino]-7-methoxyquinazolin-6-yl}oxy)piperidin-1-yl]-N-methylacetamidedi-[(2E)-but-2-enedioate] Form A

2-[4-({4-[(3-Chloro-2-fluorophenyl)amino]-7-methoxyquinazolin-6-yl}oxy)piperidin-1-yl]-N-methylacetamide(Compound (I)) (2.75 g at assumed 100% w/w, 5.80 mmol) was dissolved inethyl acetate (94 ml) and isopropanol (14 ml). The solution wasdistilled such that 25.2 ml of distillates were collected. The solutionwas cooled to 40° C. A clarified solution of fumaric acid (1.38 g, 11.90mmol) in isopropanol (21 ml) was then added over 1 hour. Compound (I)difumarate Form A seed was added (3.7 mg, 5.3 μmol). The filter used toclarify the fumaric acid solution was then washed with isopropanol (2ml). After holding for 1 hour at 40° C. the reaction was cooled to 20°C. over 2 hours. The reaction mixture was held for 15 hours beforeisolating the product by filtration. The filter cake was washed twicewith ethyl acetate (4.3 ml): isopropanol (1.2 ml) and then dried invacuo at 40° C. to give the title compound as a white solid (72.32 g,90%); ¹H NMR Spectrum: (DMSO) 1.85 (m, 1H), 2.08 (m, 1H), 2.50 (m, 1H),2.66 (d, 3H), 2.83 (m, 1H), 3.05 (s, 2H), 3.96 (s, 3H), 4.58 (m, 1H),6.64 (s, 4H), 7.23 (s, 1H), 7.28 (m, 1H), 7.46 (ddd, 1H), 7.55 (m, 1H),7.70 (broad q, 1H), 7.85 (s, 1H), 8.38 (s, 1H).

Example 5 Preparation of Compound (I) Difumarate Form A:2-[4-({4-](3-Chloro-2-fluorophenyl)amino]-7-methoxyquinazolin-6-yl}oxy)piperidin-1-yl]-N-methylacetamidedi-[(2E)-but-2-enedioate] Form A

2-[4-({4-[(3-Chloro-2-fluorophenyl)amino]-7-methoxyquinazolin-6-yl}oxy)piperidin-1-yl]-N-methylacetamide(Compound (I)) (1 g, 1.86 mmoles) and fumaric acid (0.44 g, 3.81 mmoles)were suspended in water (4.4 g) and heated to 85° C. The reactionmixture was cooled to 60° C. at 1° C./minute and Compound (I) Form Aseed was added when the temperature was 77° C. The resulting solid wasisolated by filtration, washed twice with acetone (0.70 g per wash) anddried in a vacuum oven at 40° C. to afford the title compound (0.89 g,68% yield), 1H NMR (400 MHz, DMSO-d6) d ppm 1.84 (m, 2H) 2.08 (m, 2H)2.55 (m, 2H) 2.63 (d, J=4.7 Hz, 3H) 2.86 (m, 2H) 3.12 (s, 2H) 3.93 (s,3H) 4.59 (tt, J=7.8, 3.7 Hz, 1H) 6.62 (s, 4H) 7.21 (s, 1H) 7.27 (td,J=8.1, 1.3 Hz, 1H) 7.49 (m, 2H) 7.86 (m, 2H) 8.36 (s, 1H) 9.63 (br. s.,1H).

Compound (I) Difumarate Form A Properties

Compound (I) difumarate Form A is a free flowing powder. X-ray powderdiffraction of Compound (I) difumarate (FIG. 4) indicates that thematerial is crystalline. The most prominent peaks from the XRPD patternof Form A are described hereinbefore and listed in Table 1. DifferentialScanning calorimetry shows a single melting endotherm with an onset at210.4° C. (FIG. 5). No appreciable weight losses are observed by isThermogravimetric analysis (FIG. 6). Dynamic Vapour Sorptiondemonstrates the compound to be non-hygroscopic absorbing <0.5% moistureup to 95% relative humidity with no evidence of hysteresis (FIG. 7).

Comparison of Aqueous Solubility of Compound (I) and Compound (I)Difumarate Form A

The solubility (48 hours, 25° C.) of Compound (I) in a range of aqueousbuffers is detailed in Table 4. Compound (I) exhibits pH dependentsolubility in aqueous buffers with high solubility at low pH (27.2mg/ml, pH 2.7) and low solubility at high pH (1 μg/ml, pH 7.9).Significant increases in solubility of Compound (I) occur at pH 6 andbelow. Therefore, at low pH values the dissolution rate of Compound (I)would be anticipated to be fast whereas at pH above 6 dissolution rateswould be anticipated to be slow.

The solubility (48 hours, 25° C.) of Compound (I) in water is 1 μg/ml(pH 7.0). In comparison, the solubility of Compound (I) difumarate FormA in water (48 hours) is 22.5 mg/ml (pH 3.5). The intrinsic dissolutionrates of Compound (I) difumarate Form A and Compound (I) was also beenmeasured in a range of aqueous buffers. Compound (I) difumarate Form Ahas a significantly higher intrinsic dissolution rate in pH 6.5phosphate buffer as shown in Table 6.

TABLE 6 Intrinsic Dissolution Rate of Compound (I) and Compound (I)difumarate Form A in aqueous buffers at 37° C. Intrinsic DissolutionRate (mgmin⁻¹cm⁻²)¹ Compound pH 6.5 SGF Compound (I) 0.01 19.2 Compound(I) difumarate 2.51 15.9 Form A ¹Measured using fibre-optic probe, λ =335 nm, disc size 4 mm, temp 37° C. SGF = Simulated Gastric Fluid

Pharmacokinetic Studies in Dogs to Comparing Compound (I) and Compound(I) Difumarate Form A

Both compounds were dosed to dogs as direct compression tabletscontaining 100 mg Compound (I) free base, or the equivalent thereof forthe tablets containing the di-fumarate salt) with the followingcomposition:

-   25% w/w Compound (I) (or equivalent thereof of the difumarate salt);-   10% w/w microcrystalline cellulose (Avicel 102);-   4% w/w croscarmellose sodium (AcDiSol);-   1% w/w sodium lauryl sulfate;-   Lactose to 99% w/w-   Magnesium Stearate to 100% w/w.-   Per tablet this equated to:-   151 mg Compound (I);-   60.4 mg Avicel;-   24.16 mg AcDiSol;-   6.04 mg sodium lauryl sulfate;-   356.36 mg Lactose; and-   6.04 mg Magnesium Stearate.-   Total tablet weight was 604 mg.

The performance of the solid dosage forms was evaluated in vitro priorto the in-vivo dog study commencing. Dissolution of solid dosage formsin pH 4.5 media (closest to sink conditions), at 100 mg (free baseequivalents) Compound (I) loading, showed >90% release after 45 minutes(Table 7) indicating the suitability of these dosage forms for use inthe dog PK study.

TABLE 7 % Dissolution of Compound (I) and Compound (I) Difumarate Form Ain pH 4.5 citrate buffer. % Dissolution Compound at 45 minutes Compound(I) 93.6 Compound (I) difumarate Form A 100.3

The performance of the tablet compositions administered orally to thedogs was compared to an intravenous injection of a 20mg dose of compound(I) comprising 4 mg.mL⁻¹ dissolved and made up to volume with 25% w/vhydroxypropyl-beta-cyclodextrin (HP-beta-CD) in water for injection pHadjusted to 4 using 1M HCl.

The results of the dog study are shown in Table 8.

TABLE 8 Summary of Pharmacokinetic Parameters for Compound (I) andCompound (I) difumarate Form A administered orally and Compound (I)administered intravenously in Male Beagle Dogs (n = 4, mean ± SE) Study0383KD - Compound (I) Formulation/Route (Dose) Difumarate Form A¹ Freeform¹ (equivalent to Parameter IV (20 mg) (100 mg) 100 mg free form)C_(max) (μmol/L) 9.0 ± 1.1 14.0 ± 1.7  32.6 ± 3.6 T_(max) (h) 0.2 ± 0.01.6 ± 0.8  1.1 ± 0.3 Half-life (h) 5.3 ± 0.2 8.2 ± 0.6 16.9 ± 4.0AUC₍₀₋₄₈₎ 33.4 ± 7.7  103.9 ± 25.3  190.3 ± 46.9 (μmol · h/L) Cl(ml/min/kg) 1.9 ± 0.5 — — Hbf (ml/min) 39 — — Vss (L/kg) 0.6 ± 0.1 — —Bioavailability_((0-48 h)) — 63.5 ± 7.0  112.5 ± 8.1  (%) ¹dosed asdirect compressed tablet; C_(max) = Peak plasma concentration T_(max) =Time to maximum plasma concentration. AUC = Area under curve Cl =Clearance Hbf = Hepatic blood flow Vss = Volume of distribution atsteady state

Table 8 shows that bioavailability and peak plasma concentration(C_(max)) of Compound (I) difumarate Form A are significantly betterthan those obtained for Compound (I) (113% compared to 64%bioavailability) as determined in a paired t-test is (n=4) at a 95%confidence level.

Examples 6 to 21 Crystalline Compound (I) Difumarate Forms B to Q XRPDAnalysis

XRPD patterns were collected using a Bruker D-8 Discover diffractometerand Bruker's General Detector System (GADDS, v. 4.1.20). An incidentmicrobeam of Cu Kα radiation was produced using a fine-focus tube (40kV, 40 mA), a Gael mirror, and a 0.5 mm double-pinhole collimator. Priorto the analysis, a silicon standard (NIST SRM 640c) was analysed toverify the Si 111 peak position. The sample was packed between 3 μmthick films to form a portable, disc-shaped specimen. The preparedspecimen was loaded in a holder secured to a translation stage. A videocamera and laser were used to position the area of interest to intersectthe incident beam in transmission geometry. The incident beam wasscanned and rastered to optimize orientation statistics. A beam-stop wasused to minimize air scatter from the incident beam. Diffractionpatterns were collected using a Hi-Star area detector located 15 cm fromthe sample and processed using GADDS. The intensity in the GADDS imageof the diffraction pattern was integrated using a step size of is 0.04°2θ. The integrated patterns display diffraction intensity as a functionof 2θ.

Variable Temperature—XRPD

Variable-temperature XRPD patterns (VT-XRPD) were collected using aShimadzu XRD-6000 X-ray powder diffractometer equipped with an AntonPaar HTK 1200 high-temperature stage. Prior to the analysis, a siliconstandard (NIST SRM 640c) was analysed to verify the Si 111 peakposition, and vanillin and sulfapyridine standards were analysed toverify the stage temperature. The sample was packed in a ceramic holderand analysed from 2.5 to 40° 2θ at 3°/minute (0.4 sec/0.02° step).

XRPD—Microplate Analysis

XRPD patterns were collected with a Bruker D-8 Discover diffractometerand Bruker's General Area Diffraction Detection System (GADDS, v.4.1.20). An incident beam of Cu Kα radiation was produced using afine-focus tube (40 kV, 40 mA), a Gael mirror, and a 0.5 mmdouble-pinhole collimator. The samples were positioned for analysis bysecuring the well plate to a translation stage and moving each sample tointersect the incident beam. The samples were analysed using atransmission geometry. The incident beam was scanned and rastered overthe sample during the analysis to optimize orientation statistics. Abeam-stop was used to minimize air scatter from the incident beam at lowangles. Diffraction patterns were collected using a Hi-Star areadetector located 15 cm from the sample and processed using GADDS. Theintensity in the GADDS image of the diffraction pattern was integratedusing a step size of 0.04° 2θ. The integrated patterns displaydiffraction intensity as a function of 2θ. Prior to the analysis asilicon standard was analysed to verify the Si 111 peak position.

Example 6 Preparation of Compound (I) Difumarate Form B

A slurry was prepared by adding enough Compound (I) difumarate Form A towater so that excess solid was present. The mixture was then agitated ina sealed vial at 4° C. for 7 days. Solids were isolated by vacuumfiltration and analysed. The XRPD pattern for the resulting Form B isshown in FIG. 8. The most prominent X-Ray Powder Diffraction peaks forForm B are shown in Table 9:

TABLE 9 Angle Intensity 2-Theta ° Count 4.7 362 6.8 984 7.4 977 9.0 45710.3 751 12.6 1446 13.3 473 13.6 375 14.0 656 14.3 414 14.6 377 15.8 57916.8 963 17.3 454 17.6 352 17.9 338 19.0 519 20.1 672 20.3 1165 21.1 38822.0 704 22.1 561 22.8 531 23.1 355 23.6 459 24.5 694 24.7 870 25.2 38825.5 530 25.9 497 26.2 2683 26.7 593 27.0 2034 27.4 697 27.6 788 27.8374 28.3 419 28.6 792 28.7 721 28.9 510 29.4 538 30.4 407Form B is thought to be a hydrate, possibly a tetra or penta hydrate.

Example 7 Preparation of Compound (I) Difumarate Form C

A slurry was prepared by adding enough Compound (I) difumarate Form A toIPA/water (90/10 v/v), so that excess solid was present. The mixture wasthen agitated in a sealed vial at 15° C. for 6 days. Solids wereisolated by vacuum filtration and analysed. The XRPD pattern for theresulting Form C is shown in FIG. 9. The most prominent X-Ray PowderDiffraction peaks for Form C are shown in Table 10:

TABLE 10 Angle Intensity 2-Theta ° Count 3.5 1014 5.3 1016 6.9 3144 9.11820 10.1 1648 10.7 919 11.1 917 11.9 1221 12.6 1733 13.3 1185 14.0 131614.3 1081 15.0 2958 15.6 1009 16.0 1161 17.3 886 17.6 897 18.1 941 18.3871 19.2 1791 19.5 3574 19.7 1957 20.2 1066 20.6 2120 21.0 2043 21.91661 22.6 1643 22.9 2277 23.3 1424 24.1 6452 24.5 1094 25.1 1701 25.61249 26.7 917 27.1 2265 27.8 884 28.0 874 28.6 826 29.5 2106 33.2 912Form C is believed to be a mixed hydrate/solvate form.

Example 8 Preparation of Compound (I) Difumarate Form D

A solution of Compound (I) difumarate was prepared in acetonitrile/water50/50 and an aliquot was added to a microplate well. The solvent wasevaporated and 2-propanol/water (90/10, v/v) was added to the well. Theplate was sonicated and then the to solvent was evaporated under vacuum.The XRPD pattern for the resulting Form D is shown in FIG. 10. The mostprominent X-Ray Powder Diffraction peaks for Form D are shown in Table11:

TABLE 11 Angle Intensity 2-Theta ° Count 3.3 14.1 5.1 39.1 6.9 35 8.814.9 10.2 10.5 12.0 13.6 14.0 18.9 15.0 22.5 16.2 26.3 17.1 19.1 19.021.4 20.2 47.6 21.6 39.2 24.2 85.2 25.8 61 26.7 39.6 28.1 30.3 29.8 28.1

Example 9 Preparation of Compound (I) Difumarate Form E

A solution of Compound (I) Difumarate was prepared in acetonitrile/water50/50 (v/v) and an aliquot was added to a microplate well. The solventwas evaporated and tetrahydrofuran was added to the well. The plate wassonicated and then the solvent was evaporated under vacuum. The XRPDpattern for the resulting Form E is shown in FIG. 11. The most prominentX-Ray Powder Diffraction peaks for Form E are shown in Table 12:

TABLE 12 Angle Intensity 2-Theta ° Count 5 1.12 6.7 13.3 8.7 26.7 10.413.3 14.2 28.7 17.0 26.4 20.5 61.5 22.9 31.4 23.8 39.8 25.2 26 26.1 57.527.3 23.2

Example 10 Preparation of Compound (I) Difumarate Form F

A saturated solution of Compound (I) difumarate was prepared intetrahydrofuran at elevated temperature and filtered through a 0.2 μmnylon filter into a pre-warmed vials while still warm. The vials werecovered and allowed to cool slowly to room temperature. The presence orabsence of solids was noted. If there were no solids present, or if theamount of solids was judged too small for XRPD analysis, the vial wasplaced in a refrigerator. Again, the presence or absence of solids wasnoted and if there were none, the vial was placed in a freezer. Solidsthat formed were isolated by filtration and allowed to dry prior toanalysis. The XRPD pattern for the resulting Form F is shown in FIG. 12.The most prominent X-Ray Powder Diffraction peaks for Form F are shownin Table 13.

TABLE 13 Angle Intensity 2-Theta ° Count 6.3 1491 8.0 1591 13.5 176413.9 1759 15.0 1349 16.0 1301 19.1 1743 20.1 1547 21.5 1327 22.8 268323.9 1397 24.8 1295 26.2 1276 27.4 970

Example 11 Preparation of Compound (I) Difumarate Form G

A solution of Compound (I) difumarate was prepared in acetonitrile/water50/50 and an aliquot was added to a microplate well. The solvent wasevaporated and 2-propanol/water 90/10, was added to the well. The platewas sonicated and then the solvent was evaporated under vacuum. The XRPDpattern for the resulting Form G is shown in FIG. 13. The most prominentX-Ray Powder Diffraction peaks for Form G are shown in Table 14.

TABLE 14 Angle Intensity 2-Theta ° Count 6.9 919 8.7 446 10.1 259 12.0488 12.8 394 14.0 223 14.9 695 16.2 244 19.0 366 20.2 1461 21.6 925 23.2848 24.1 2532 25.4 753 26.7 579 27.4 756 29.7 749

Example 12 Preparation of Compound (I) Difumarate Form H

A solution of Compound (I) difumarate was prepared in acetone/water(95/5, v/v), and sonicated between aliquot additions to assist indissolution. Once the mixture reached complete dissolution, as judged byvisual observation, the solution was filtered through a 0.2 μm nylonfilter. The filtered solution was allowed to evaporate under ambientconditions in an uncapped vial. The solids that formed were isolated andanalysed. The XRPD pattern for the resulting Form H is shown in FIG. 14.The most prominent X-Ray Powder Diffraction peaks for Form H are shownin Table 15:

TABLE 15 Angle Intensity 2-Theta ° Count 4.8 1837 5.2 2010 6.8 1639 13.21563 13.8 1603 15.8 1625 16.5 1658 16.9 1603 23.3 1786 25.2 2564 25.42231 26.1 1698 27.6 1864

Example 13 Preparation of Compound (I) Difumarate Form I

A solution of Compound (I) difumarate was prepared in methanol atambient is temperature. The solution was then filtered into toluene atambient temperature. The resulting solid was isolated by filtration anddried prior to analysis. The XRPD pattern for the resulting Form I isshown in FIG. 15. The most prominent X-Ray Powder Diffraction peaks forForm I are shown in Table 16:

TABLE 16 Angle Intensity 2-Theta ° Count 10.7 2170 11.1 1793 14.3 172814.8 2080 16.0 2165 17.4 1868 21.7 1947 22.1 2130 24.8 1688 26.8 1643

Example 14 Preparation of Compound (I) Difumarate Form J

A solution of Compound (I) difumarate was prepared in methanol atambient temperature. The solution was then filtered into an excess ofheptane at ambient temperature. The resulting solid was isolated byfiltration and dried prior to analysis. The XRPD pattern for theresulting Form J is shown in FIG. 16. The most prominent X-Ray PowderDiffraction peaks for Form J are shown in Table 17:

TABLE 17 Angle Intensity 2-Theta ° Count 7.0 1804 7.6 2353 8.7 2306 10.52129 11.9 1719 12.2 1802 12.9 1696 14.7 2171 16.1 1728 17.4 2079 19.12165 20.1 2723 21.6 3417 22.5 2094 23.8 3119 24.4 2048 25.4 2041 27.31978 29.5 1501

Example 15 Preparation of Compound (I) Difumarate Form K

A solution of Compound (I) difumarate was prepared in acetonitrile/water(50/50 v/v) and an aliquot was added to a microplate well. The solventwas evaporated and fluorobenzene was added to the well. The plate wassonicated and then the solvent was vaporated under ambient conditions.The XRPD pattern for the resulting Form K is shown in FIG. 17. The mostprominent X-Ray Powder Diffraction peaks for Form K are shown in Table18:

TABLE 18 Angle Intensity 2-Theta ° Count 10.5 32.3 11.8 109 12.3 96.713.0 49.4 14.2 11.8 16.1 41 20.4 137 21.4 128 21.8 112 23.0 76.3 23.9101 24.6 198 25.5 133 27.4 191 29.4 140 31.8 39.3 33.9 50.6 34.8 69.6

Example 16 Preparation of Compound (I) Difumarate Form L

A solution of Compound (I) difumarate was prepared in acetonitrile/water50/50 (v/v) and an aliquot was added to a microplate well. The solventwas evaporated and 1,1,1,3,3,3 hexafluoro-2-propanol was added to thewell. The plate was sonicated and then the solvent was evaporated underambient conditions. The XRPD pattern for the resulting Form L is shownin FIG. 18. The most prominent X-Ray Powder Diffraction peaks for Form Lare shown in Table 19:

TABLE 19 Angle Intensity 2-Theta ° Count 7.1 13.7 9.7 10.1 11.1 17.914.7 17 18.2 20.4 18.7 15.7 19.6 14.8 21.2 25.8 21.8 21.1 22.9 55.4 23.521.8 26.0 34.7 26.9 30.4 27.7 24.1 28.9 42.7 29.5 19.8 33.7 11.4

Example 17 Preparation of Compound (I) Difumarate Form M

A solution of Compound (I) difumarate was prepared in acetonitrile/water(50/50 v/v) and an aliquot was added to a microplate well. The solventwas evaporated and 2-propanol/water (90/10 v/v), was added to the well.The plate was sonicated and then the solvent was evaporated underambient conditions. The XRPD pattern for the resulting Form M is shownin FIG. 19. The most prominent X-Ray Powder Diffraction peaks for Form Mare shown in Table 20:

TABLE 20 Angle Intensity 2-Theta ° Count 5.3 41.8 6.7 18.6 16.1 20 20.1125 21.5 63.3 23.9 72.2 25.4 54.8 27.4 41.9 29.5 25.8 31.2 25.4

Example 18 Preparation of Compound (I) Difumarate Form N

A solution of Compound (I) difumarate was prepared in acetonitrile/water(50/50 v/v) and an aliquot was added to a microplate well. The solventwas evaporated and acetone/water (60/40 v/v), was added to the well. Theplate was sonicated and then the solvent was evaporated at 4° C. TheXRPD pattern for the resulting Form N is shown in FIG. 20. The mostprominent X-Ray Powder Diffraction peaks for Form N are shown in Table21:

TABLE 21 Angle Intensity 2-Theta ° Count 5.3 59.2 6.9 36.2 9.2 23.2 10.58.5 12.2 9.01 13.3 25.1 13.9 28.2 23.5 51.1 25.6 80.3 26.4 60.5 27.862.3

Example 19 Preparation of Compound (I) Difumarate Form O

A solution of Compound (I) difumarate was prepared in acetonitrile/water(50/50 v/v) and an aliquot was added to a microplate well. The solventwas evaporated and ethanol/water (30/70 v/v), was added to the well. Theplate was sonicated and then the solvent was evaporated at 4° C. TheXRPD pattern for the resulting Form O is shown in FIG. 21. The mostprominent X-Ray Powder Diffraction peaks for Form O are shown in Table22:

TABLE 22 Angle Intensity 2-Theta ° Count 6.9 28.5 8.7 21.7 11.9 27.614.9 36.8 20.2 87.4 21.8 70.6 24.0 223 25.5 59.1 26.4 56.2 27.4 63.729.5 58.4

Example 20 Preparation of Compound (I) Difumarate Form P

A solution of Compound (I) difumarate was prepared in acetonitrile/water(50/50 v/v) and an aliquot was added to a microplate well. The solventwas evaporated and 2-propanol/water (90/10 v/v) was added to the well.The plate was sonicated and then the solvent was evaporated at 4° C. TheXRPD pattern for the resulting Form P is shown in FIG. 22. The mostprominent X-Ray Powder Diffraction peaks for Form P are shown in Table23:

TABLE 23 Angle Intensity 2-Theta ° Count 5.3 9.97 7.2 9.97 14.8 15.824.1 30.6 25.6 29.3 26.5 23.2 27.2 27.9 27.9 20.3 29.5 23.2

Example 21 Preparation of Compound (I) Difumarate Form Q

Form Q was observed when Form B was heated to 150° C. during variabletemperature XRPD analysis. The XRPD pattern for Form Q is shown in FIG.23. The most prominent X-Ray Powder Diffraction peaks for Form Q areshown in Table 24:

TABLE 24 Angle Intensity 2-Theta ° Count 8.0 92 12.5 210 17.8 130 18.9212 20.9 318 22.7 316 23.9 272 24.9 290 25.9 358 26.3 280 27.6 110 35.4552 38.1 244

Example 22 Tablet formulation of Compound (I) Difumarate

The powdered ingredients shown below were charged to a mixer and mixedto produce a uniform distribution of Compound (I) difumarate. A bindersolution was prepared and added to the powders with further mixing untila suitable wet mass formed. The wet mass was passed through a screen andthe resultant granules dried to an appropriate moisture content (forexample less than 2% by weight). The dried granules were passed throughan appropriately sized screen and blended with magnesium stearate beforecompressing into tablet cores using conventional tabletting equipment.The compressed cores were then coated with an aqueous suspension of filmcoating components using a conventional perforated drum coater.

Film-coated tablets containing 2.5, 10, 40 and 100 mg of Compound (I)Difumarate Form A prepared as described above are illustrated in Table25.

TABLE 25 Tablet strength¹ 2.5 mg 10 mg 40 mg 100 mg Ingredient g/batchg/batch g/batch g/batch Tablet core Compound (I) 37.25 149.0 448.1 448.1Difumarate Form A² Lactose (450 mesh) 782.75 671.0 371.9 371.9Microcrystalline 100.0 100.0 100.0 100.0 cellulose (PH101) Crospovidone50.0 50.0 50.0 50.0 Polyvidone 20.0 20.0 20.0 20.0 Magnesium stearate10.0 10.0 10.0 10.0 Core tablet 100 mg 100 mg 133 mg 333 mg weightTablet coating Opadry White 23.0 23.0 23.3 23.0 (03B28460) Hypromellose³15.0 15.0 15.0 15.0 Titanium dioxide³ 5.0 5.0 5.3 5.0 Macrogol 300³ 3.03.0 3.0 3.0 Purified water⁴ 177.0 177.0 176.7 177.0 Nominal coatedtablet 102.1 mg 102.1 mg 136.1 mg 140.6 mg weight ¹Tablet strengthsrefer to the equivalent amount of Compound (I) free base present in thetablet. ²The Compound (I) difumarate was micronised prior to formulationto give an average particle size of less than about 5 μm. ³Thehypromellose, macrogol 300 and titanium dioxide are included as OpadryWhite (03B28460), supplied by Colorcon. ⁴Purified water is used as thesolvent/carrier fluid during film-coating and is removed during thecoating process.A suitable manufacturing process is outlined below:

1.4-(3-chloro-2-fluoroanilino)-7-methoxy-6-{[1-(N-methylcarbamoylmethyl)piperidin-4-yl]oxy}quinazolinedifumarate Form A.